For treating the fit modified polymeric material of combination of the factor in wound climateThis application claims the priority of the U.S.Provisional Serial 61/518,148 submitted on April 29th, 2011,The entire disclosure of which is incorporated herein by reference.
1. invention field
The present disclosure generally relates to wound healing and treatment of wounds therapy.More specifically, but be not with limit sideFormula, present disclosure is related to modified material, for example, the silyl-modified poly- ammonia of the Reversible binding for the factor in wound bedEster foam.
2. background technology
Clinical research and practice are promoted and are accelerated in the group it has been shown that being provided about decompression in a tissue siteKnit the neoblastic growth at position.The application of this phenomenon is a lot, but it is particularly successful in terms of wound is processed to apply decompression.This process (being frequently termed as " negative pressure wound therapy ", " reduced pressure therapy " or " vacuum therapy " in medical circle) provides substantial amounts ofAdvantage, including the generation of the granulation tissue for faster healing and increasing.One of Major Clinical benefit of negative pressure wound therapyBe it effectively eliminate Wound exudate, so as to reduce edema and allow tissue reduce pressure ability.Negative pressure wound therapy mayIt is not always able to distinguish the harmful and beneficial agents removed from wound.For solve the coating of this point for example collagen, PVA,PEG and Fibrinogen Jing often has the shortcomings that it is not covalent, uneven or non-target-specific.Typically, theyCan not be presented in the way of allowing active site to retain its function in a specific part conjugated protein and by themCell.Therefore, molecule will be allowed to be combined in a kind of covalent mode, it is stipulated that the type of combinable compound, combinationTheir chemical reaction, and/or by protein be presented to cell orientation (for example, for a kind of deposited part, wound insert,Pad etc. used in) improvement will be very desirable.
General introduction
The present disclosure provides can be used for the new material of the combination of the factor (such as from a wound bed), including it is fit(for example, polypeptide) modified polymer.In certain embodiments, these factors are endogenic.In other embodiments, theyIt is ectogenic.In certain embodiments, the combination is reversible, and in other embodiments, it is irreversible.
In certain embodiments, there is provided a kind of polypeptide, the polypeptide is included and SEQ ID NO:2(P16；GHQGQHIGQMS)An aminoacid sequence unanimously and SEQ ID NO:A 2 at least 90% consistent sequences, or relative to SEQ ID NO:2A sequence comprising 1,2 or 3 aminoacid replacement or disappearance.In some respects, the aminoacid sequence further includes oneAmino or carboxyl terminal Cys residues (for example, a kind of amino or carboxyl terminal of polypeptide of these embodiments comprising PEG interval bases-Cysteine residues).For example, the aminoacid sequence can include sequence AEEAc-Cys-NH in the C- ends2.Still anotherOuter aspect, there is provided a kind of wound applies part, and the wound applies part includes that a kind of one kind of conjugated polypeptide to these embodiments is gatheredCompound foam substrate, the polypeptide is as included SEQ ID NO:2 (GHQGQHIGQMS) aminoacid sequence or with SEQ ID NO:A kind of polypeptide of a 2 at least 90% consistent sequences.
In certain embodiments, there is provided a kind of wound applies part, and it is poly- including conjugated to a kind of one kind of polypeptide that the wound applies partCompound foam substrate, the wherein polypeptide are included and SEQ ID NO:1(P22；NAIQEARRLLNLSRD) consistent aminoacidSequence and SEQ ID NO:A l at least 90% consistent sequence, or relative to SEQ ID NO:1 includes 1,2 or 3 aminoThe sequence that acid replaces or lacks.In some respects, the aminoacid sequence is further comprising an amino or carboxyl terminal CysResidue (for example, a kind of amino or carboxyl terminal of polypeptide of these embodiments includes PEG interval bases-cysteine residues).ExampleSuch as, the aminoacid sequence can include sequence AEEAc-Cys-NH in the C- ends2。
In certain embodiments, a kind of polypeptid covalence of these embodiments be attached to a kind of foam of polymers (for example, so as toForm a kind of wound and apply part).In some respects, a kind of polypeptide is attached to the foam of polymers by a thioether linker.ExampleSuch as, the linker can be linker derived from an EMCS or linker derived from a sulfo group-EMCS.In some respects,The thioether linker includes a Cys residue in the polypeptide, such as an AEEAc-Cys-NH of the polypeptide2One in sequenceCys residues.A linker between still other aspect, a kind of polypeptide and a kind of foam of polymers can be a firstLinker derived from silylation is (for example, derived from amino-undecanoic ethyl triethoxy silicane alkane or aminopropyl diisopropyl ethoxyThe silicyl of one replacement of base silane).At still other aspect, a kind of polypeptide of these embodiments is via a kind of non-co-Valency combines (as by a kind of biotin-avidin combination) and is attached to a kind of foam of polymers.
In some aspects, a kind of wound of these embodiments applies part further comprising one kind life for being bound to the deposited part of the woundThe long factor, chemotactic factor or cytokine (for example, are bound to the wound via the interaction with the polypeptide of these embodimentsDeposited part).In some respects, the somatomedin is granulocyte macrophage colony stimulating factor (GM-CSF).In some aspects, shouldSomatomedin is VEGF (VEGF).
In certain embodiments, a kind of polypeptide of these embodiments is attached to a kind of foam of polymers, so as to form one kindWound applies part.For example, the foam of polymers substrate can be a kind of netted open-celled foam.The example bag of foam of polymersInclude, but be not limited to, the foam substrate comprising following thing：Polyvinyl alcohol, polyurethane, polypropylene, polystyrene, polyhydric alcohol, poolLuo Shamu, aminoglycoside, amino sugar or one of these or multiple combinations.
In a further embodiment, there is provided a kind of method for binding growth factor, chemotactic factor or cytokine,The method is included by a kind of fluid (such as body fluid) comprising the somatomedin, chemotactic factor or cytokine and according to embodimentWound applies part and contacts, so as to reference to the somatomedin, chemotactic factor or cytokine.In some respects, these embodimentsA kind of wound applies part and is adapted to apply negative pressure to a wound location.In a further aspect, a kind of method includes removingSome or all in the fluid that part contacts are applied with the wound.In still other aspect, a kind of method of these embodimentsFurther include that the wound is applied into part contacts with least one second fluid, wherein be bound to the growth of the deposited part of the wound becauseSome or all in son, chemotactic factor or cytokine are eluted among the second fluid.In some aspects, these enforcementsA kind of method of example is defined as a kind of in vitro method.
In further embodiment again, there is provided a kind of compositionss for processing wound, said composition is included according to thisA kind of wound of a little embodiments applies part.In some respects, there is provided a kind of method for processing wound, the method is included oneIndividual wound location is applied part and is contacted with a kind of wound of these embodiments.In some aspects, a kind of method of these embodiments is enteredOne step includes applying negative pressure to the wound location.
Arbitrarily any embodiment of the system of the present invention and/or method can by or substantially by arbitrarily described stepSuddenly, element and/or feature composition (rather than comprising/include/containing/there is arbitrarily described step, element and/or spyLevy).Therefore, in any claim, term " by ... constitute " or " substantially by ... constitute " above-mentioned can be replacedMeaning open copulative verb, to change the scope of given claim, otherwise described scope be using it is open link it is dynamicWord by for scope.
Other targets of present disclosure, feature and advantage will be made apparent from from described further below.It will be appreciated, however, thatAlthough this detailed description and these instantiations indicate the specific embodiment of the present invention, but they are merely by explanationWhat mode was given, because the difference from this detailed description within the spirit and scope of the present invention is changed and modifications for this areaThose those of ordinary skill for will become apparent from.It should be noted that merely because a kind of particular compound belongs to one specificallyFormula is not meant to that it can not still belong to another formula.
Brief Description Of Drawings
By way of example rather than restrictive one illustrates the following drawings.It is given manifesting in order to simple and clear and clearStructure per width figure in each feature without the always labelling given structure.Same reference numbers are unnecessary to indicate identical knotStructure.But, same reference numbers can serve to indicate that similar characteristics or the feature with similar functions, differ reference number sameSample can be such.
Fig. 1 depicts the side view that the wound of the present invention applies one embodiment of part, and the wound applies part and has the present invention'sOne kind in modified wound insert and it is attached to a wound location and a Wound treating apparatus.
Fig. 2 depicts the enlarged side view of the modified wound insert of Fig. 2.
Fig. 3 depicts the block schematic diagram of one embodiment of a Wound treating apparatus, and the Wound treating apparatus can be withIncluding and/or be attached to and/or apply part and/or modified wound insert with the wound of the present invention and be used together.
Fig. 4 depicts the schematic diagram of some embodiments of the invention.In this side view, one kind has free hydroxylBase group, based on a kind of R ' of the polymer of polyurethane-(CH2)n-Si(OH)3Reagent is modified.
Fig. 5 is depicted to be determined using o-phthalaldehyde(OPA) (OPA) and is carried out the silylated 30phr shitosans foam of comparisonThe result of (" 30PHR Si ") and the number of the free amino group of silylated reticulated, open type foam (" ROCF ").At two kindsIn the case of, silylating group is from the (H of compound 602 (aminopropyl diisopropyl Ethoxysilane)2N(CH2)3)(i-Pr)2Si-.These results are providing according to Relative fluorescence units (RFU).OPA is individually for the sensitive of amineDetectable, it also serves as control.
Fig. 6 depicts the group (H compared with from compound 630 (amino-undecanoic ethyl triethoxy silicane alkane)2N(CH2)11)(EtO)2Si- silylanizings and use TexasThe reticulated, open type foam of process and non-silylanizingReticulated, open type foam (" ROCF ") (individually and and TexasThe knot of Relative fluorescence units (" RFU ") together)Really.These results are provided according to Relative fluorescence units (RFU).Also analyze the ROCF in PBS and PBS to guarantee these materialsThe autofluorescence of material will not make RFU reading offsets.PBS is phosphate buffered saline (PBS).
Fig. 7 is depicted based on toluene di-isocyanate(TDI) (TDI), shitosan and VoranolTMThe polymerization of 3010 trihydroxylic alcoholsA kind of polyurethane three repetitives.
Fig. 8：Silylanizing is carried out with compound 602, it is thin that 30PHR shitosans/P-22 captures more granulocyte macrophagesBorn of the same parents' colony stimulating factor (GM-CSF).Briefly, the GM-CSF of 333ng/mL is incubated into 24 hours together with these samples.WillUnconjugated protein is washed off, then these samples is carried out into eluting with a kind of buffer (E1) based on imidazoles.By these samplesProduct carry out buffer-exchanged to be purified via with PBS, and are quantified by ELISA.As a result indicate with P-22Positive captures the GM-CSF of 299pg/mL, and 30PHR individually combines 124pg/mL.
Fig. 9 A and Fig. 9 B provides the ROCF of silanization and the image of texas Red (Texas Red) fluorescent dye.Fig. 9 AOne bright vision image is shown, the image illustrates the orientations and form of-Sil/ROCF in normal light.Fig. 9 B illustrate it is conjugated extremelyThe TRITC imagings of 630 texas Red, the imaging illustrates that the dyestuff is conjugated with the success of-Sil.This provides 630 successfullyThe evidence being deposited on the ROCF.Two images are obtained under 20 × amplification.
Figure 10 provides the structure and connectivity information of a part for a fit modified foam of polymers embodiment, and this is fittedModifies foam of polymers embodiment includes a kind of oligopeptide (P16；SEQ ID NO:2), AEEAc-Cys linker, oneIndividual EMCS linkers, one of the silicyl of replacement and the trihydroxylic alcohol based on PEGization and toluene di-isocyanate(TDI) it is poly-Urethane main chain.
Figure 11 provides the P22/GM-CSF eluting results of experiment.P22 is from Bach's nurse u s company (BachemAmericas Inc.) 2010 peptide catalogues obtain.P22 is the GM-CSF antagonisms for combining GM-CSF in a kind of inhibition modeAgent.Briefly, by 330nmol P22 via conjugated to a kind of ROCF based on polyurethane of sulfo group-EMCS chemistry.By the P22-Linker-foam construct overnight incubation, washing five times and undergoes strict eluting together with 1 μ g GM-CSF.Before elutingWashing show the protein that non-specifically combines be reduced to insignificant level (<100pg/mL).In eluting, haveThe at most sample S3 of capture peptide is more than ROCF individually up to 49%.The total amount (608pg/mL) for being bound to the GM-CSF of ROCF/P22 is bigIn the amount (376pg/mL) of the GM-CSF for being non-specifically bound to negative control C1.Sample S1 to S3 all comprising peptide-linker-Foam, wherein sulfo group-EMCS concentration increase (S1=10 × sulfo group-EMCS+P22 from S1 to S3；S2=25 × sulfo group-EMCS+P22 and S3=50 × sulfo group-EMCS+P22), C1 be ROCF individually, C2 is the ROCF+EMCS-P22 of silanization, and C3 isThe ROCF-EMCS+P22 of silanization.The positive control of this experiment it is not available for, because also proving to be total to peptide without technologyValency ground is conjugated to polyurethane.However, being based on experimental result, positive control is developed afterwards is used for subsequent experiment.Data are used as flatMean value ± standard deviation is given.
Figure 12 provides the structure and connectivity information of a part for a fit modified foam of polymers embodiment, and this is fittedModifies foam of polymers embodiment includes a kind of oligopeptide (P16；SEQ ID NO:2), AEEAc-Cys linker, oneIndividual EMCS linkers, a silicyl for replacing, and it is poly- based on the trihydroxylic alcohol of PEGization, toluene di-isocyanate(TDI) and shellA kind of polyurethane-copolymer of sugared oligomer.
Figure 13 provides the structure and connectivity information of a part for a fit modified foam of polymers embodiment, and this is fittedModifies foam of polymers embodiment includes a kind of oligopeptide (P16；SEQ ID NO:2), AEEAc-Cys linker, oneIndividual EMCS linkers, and a kind of poly- ammonia based on the trihydroxylic alcohol of PEGization, toluene di-isocyanate(TDI) and shitosan oligomerEster-copolymer.
Figure 14 illustrates ROCF in the case where it is covalently bond to P16 (for the capture peptide for from the beginning designing of VEGF)The VEGF of capture more than 82%.C1 is ROCF independent；C2 is ROCF+EMCS-P16；C3 is ROCF-EMCS+P16；C4 is silaneThe ROCF of change is independent；The ROCF+EMCS-P16 of C5=silanizations；C6 is the ROCF-EMCS+P16 of silanization；S1 is 50 × sulphurBase-EMCS+P16；And S2 is 100 × sulfo group-EMCS+P16.W=final follow-up PBS washings.E1=50mM Tris,80mM NaCl, 250mM imidazoles, 45 minutes.W5 (left side) is illustrated and is non-specifically retained in ROCF in final PBS washingsVEGF(pg/mL).E1 (right side) indicates the VEGF removed from the foam and peptide with a kind of organic solvent after last time is washedAmount.Elution profile indicates that S2 (P16- linkers-foam) (has the VEGF of most abundant combination than C4 (ROCF of silanization)Negative control) combine 82% VEGF.S2 combines 584pg/mL, and C4 combines 321pg/m.S2 has maximum concentrationEMCS and therefore at most covalently bound P16.This data display is covalently attached to the amount of the P16 of foam and is caught by the foamStrong correlation between the amount of the target protein caught.Data are given as meansigma methodss ± standard deviation.
Figure 15 illustrates a kind of fit modified in a kind of deposited part for being used in combination with negative pressure wound therapy (NPWT)Polymer.In this embodiment, this is fit selective for MCP1 (the chemotaxiss molecule of macrophage).It is this modifiedDeposited part can be used for stimulating expression of macrophage and migrate into wound and thus make the wound advance to healing status from chronic.
Figure 16 provides the structure and connectivity information of a part for a fit modified foam of polymers embodiment, and this is fittedModifies foam of polymers embodiment is included and is covalently attached to based on the trihydroxylic alcohol of PEGization, toluene di-isocyanate(TDI) and shellA kind of a kind of oligopeptide (P16 of the polyurethane-copolymer of polysaccharide oligomer；SEQ ID NO:2).
Figure 17 depicts the schematic diagram of some embodiments of the invention.In this side view, with specially designedA kind of protein-modified insert is shown with different dissociation constant (KdValue) it is bound to the different factors.In this embodiment,The modified insert is used containing plurality of reagents (for example, for the examination of wound clean, promotion healing, stability of solution etc.Agent) processing solution (the bolus agent (bolus) of metering) be rinsed (continuously or intermittently).Block arrow (blockArrow) " exudate " of labelling represent and leave the mixture of the material, the mixture comprising processing solution and from may with applyThe exudate of the wound that part contacts.
Figure 18 --- the VEGF in solution is made by being connected to a kind of beadlet of VEGF antibody.Pass through this solution is madeAfter a little beadlet, centrifugation (spun down) and these beadlet are washed.The beadlet of these washings is subsequently used for endotheliocyteMigration is determined.The result of this experiment shows after 3 hours, compared with the beadlet without antibody, 2 times more of cell migration courtTo these beadlet for being bound to the antibody (and VEGF).
The explanation of illustrative embodiment
In some respects, present disclosure provides fit modified material, and in certain embodiments, these materials can serve as usingIn the wound for catching and concentrating the specificity factor (for example, causing a kind of factor of biological respinse such as angiogenesis) in wound climateInsert, or for biomolecule or bioactive compound to be delivered to into a wound bed.In certain embodiments, these becauseSon is endogenic.In other embodiments, they are ectogenic.In certain embodiments, the combination is reversible, at itIn his embodiment, it is irreversible.
In one aspect, the present invention provides the deposited part of wound of fit functionalization, and these wounds apply part to be included：(a) liningBottom；And (b) is covalently attached to a kind of fit of the foam of polymers.In certain embodiments, the substrate is a kind of polymerFoam.In certain embodiments, this is fit comprising a kind of polypeptide, a kind of small molecule binding agent, a kind of dendritic, one kindNano-particle, and/or a kind of oligonucleotide.In certain embodiments, it is described it is fit be a kind of polypeptide.In certain embodiments,The polypeptide is synthesis.In certain embodiments, the polypeptide is natural.
In certain embodiments, the polypeptid covalence is attached to the foam of polymers.In certain embodiments, the C of the polypeptideEnd includes PEG interval bases-cysteine residues.In certain embodiments, this it is fit by a linker be attached to this gatherCompound foam.In certain embodiments, the linker is connected derived from linker derived from an EMCS or a sulfo group-EMCSConnect base.In certain embodiments, the linker is linker derived from a silicyl for replacing.In certain embodiments,Linker derived from the substituted silicyl is derived from compound 630 (amino-undecanoic ethyl triethoxy silicane alkane).OneIn a little embodiments, linker derived from the substituted silicyl is derived from (the aminopropyl diisopropyl ethoxy of compound 602Base silane).In certain embodiments, the linker includes first and second linkers, and wherein first linker is oneLinker derived from individual EMCS or linker derived from a sulfo group-EMCS, and second linker is the first of a replacementLinker derived from silylation.
In certain embodiments, the linker is bound to the foam of polymers by an oxygen atom.In some embodimentsIn, the linker is bound to the foam of polymers by a nitrogen-atoms.In certain embodiments, this it is fit be configured to combineEndogenouss or exogenous factor into wound climate.In certain embodiments, the combination is irreversible.In some embodimentsIn, the combination is reversible.In certain embodiments, this fit is configured to be bound to VEGF.At someIn embodiment, this is fit to be configured to be bound to platelet-derived somatomedin.In certain embodiments, this fit is configuredInto being bound to fibroblast growth factor.In certain embodiments, this fit is configured to be bound to keratinocyte growthThe factor.In certain embodiments, this fit is configured to be bound to granulocyte macrophage colony stimulating factor.
In certain embodiments, the substrate is a kind of reticulated, open type foam.In certain embodiments, the substrate is comprising poly-Vinyl alcohol, polyurethane, polypropylene, polystyrene, polyhydric alcohol, poloxamer, aminoglycoside, amino sugar or one of theseOr multiple combinations.In certain embodiments, the foam of polymers includes a kind of first monomer subunits.In some embodimentsIn, first monomer subunits are a kind of diisocyanate.In certain embodiments, the diisocyanate is toluene diisocynateEster, methylenediphenyl diisocyanates or the ethyl isocyanate of lysine two.In certain embodiments, the foam of polymers entersOne step includes a kind of second comonomer subunit.In certain embodiments, the second comonomer is a kind of trihydroxylic alcohol.In some embodimentsIn, the trihydroxylic alcohol is a kind of glycerol of PEGization.In certain embodiments, the foam of polymers is further single comprising a kind of 3rdBody subunit.In certain embodiments, the Third monomer subunit is a kind of aminoglycoside.In certain embodiments, the aminoGlucosides is shitosan.In certain embodiments, the aminoglycoside is glycosamine.In certain embodiments, the Third monomer is sub- singleUnit is neomycin (neomycin).In certain embodiments, the Third monomer subunit is dibekacin (dibekacin).In some embodiments, the Third monomer subunit is kanamycin (kanamycin).In certain embodiments, the Third monomerSubunit is tobramycin (tobramycin).In certain embodiments, the Third monomer subunit is streptomycin(streptomycin).In certain embodiments, the Third monomer subunit is gentamycin (gentamicin).In some realitiesIn applying example, the foam of polymers further includes a kind of oligomer subunit.In certain embodiments, the oligomer subunit isA kind of oligomer based on shitosan.In certain embodiments, the oligomer subunit is a kind of oligomer based on glycosamine.
On the other hand, the present invention provides fit modified polymer, and wherein the main polymer chain is comprising with followingizationOne the first repetitive of formula：
Wherein：
U1 is the unmodified part of first repetitives；
L1It is a linker molecule or a key；And
L2It is a linker molecule or a key.
In certain embodiments, L1 is that the chemical formula of a key and first repetitives is further defined as：
In certain embodiments, L2 is that the chemical formula of a key and first repetitives is further defined as：
In certain embodiments, L1 and L2 are that the chemical formula of key and first repetitives is further defined as：
In certain embodiments, U1 has below formula：
The attachment point of the wherein side chain is linked to L1。
In certain embodiments, U1 has below formula：
The attachment point of the wherein side chain is linked to L1。
In certain embodiments, U1 has below formula：
The attachment point of the wherein side chain is linked to L1.
In certain embodiments, U1 has below formula：
Wherein：
N1, n2 and n3 are independently of one another from 0 to 20；And
The attachment point of the side chain is linked to L1。
In certain embodiments, U1 is based on a kind of monomer based on trihydroxylic alcohol.In certain embodiments, ternary should be based onThe monomer of alcohol is a kind of glycerol molecule of PEGization with the molecular weight between 500 and 5,000g/ mole.
In certain embodiments, L1With chemical formula-O-Si (R1)(R2)-R3- X-, wherein：
R1And R2It is independently hydrogen, hydroxyl, halogen, alkyl(C≤6)Or alkoxyl(C≤6)；
R3It is alkane diyl(C≤20)；And
X is-NH- ,-C (O)-or-S-；And wherein X is linked to L2Or to be linked to this fit.
In certain embodiments, R1And R2It is isopropyl.In certain embodiments, R1And R2It is hydroxyl.In some embodimentsIn, R3It is alkane diyl(C3-11).In certain embodiments, R3It is-(CH2)3-.In certain embodiments, R3It is-(CH2)11-.OneIn a little embodiments, X is-NH-.
In certain embodiments, L1Or L2It is a kind of amino sulfenyl linker.In certain embodiments, L1Or L2It is a kind ofNHS- maleimide crossing linking reagents.In certain embodiments, L1Or L2It is a kind of NHS- haloacetyls based cross-linker.In some realitiesIn applying example, L1Or L2It is a kind of NHS- pyridyldithiols cross-linking agent.
In certain embodiments, derived from a kind of reagent being selected from the group, the group is by following for the amino sulfenyl linkerItem composition：SM (PEG) 24, SM (PEG) 12, SM (PEG) 8, SM (PEG) 6, SM (PEG) 4, sulfo group-LC-SMPT, SM (PEG) 2,Sulfo group-KMUS, LC-SMCC, LC-SPDP, sulfo group-LC-SPDP, SMPH, sulfo group-SMPB, SMPB, SMPT, SIAB, sulfo group-SIAB, sulfo group-EMCS, EMCS, sulfo group-SMCC, SMCC, MBS, GMBS, sulfo group-GMBS, sulfo group-MBS, SPDP, SBAP,BMPS, AMAS and SIA.
In certain embodiments, the fit modified polymer includes a kind of second repetitives.In certain embodiments,Second repetitives are a kind of oligomer based on shitosan.
In certain embodiments, U1 has below formula：
The attachment point of the wherein side chain is linked to L1.
In certain embodiments, L1 has below formula：
In certain embodiments, L2 has below formula：
In certain embodiments, there is L2 chemical formula-C (O) (CH2) 5-S-Cys-AEEAc-, wherein AEEAc to be：
In certain embodiments, the fit modified polymer further includes a kind of second repetitives.
In certain embodiments, second repetitives have below formula：
In certain embodiments, fit is a kind of oligopeptide with 8 to 12 residues.In some are example, the oligopeptideWith sequence GHQGQHIGQMS.In certain embodiments, fit is P16.In certain embodiments, fit is P22.
In certain embodiments, the fit modified polymer is further triple comprising the one kind the with below formulaMultiple unit：
Wherein X is：
- alkane diyl(C≤12)-；
- fragrant diyl(≤12)-；
- fragrant diyl(≤6)-CH2- fragrant diyl(≤6)-；Or
The substituted pattern of any these groups.
In certain embodiments, X is further defined as：
In certain embodiments, the third repeating unit is a polyurethane repetitives.In certain embodiments, the 3rdRepetitives are the polyurethane repetitives based on ether.In certain embodiments, the third repeating unit is one and is based onThe polyurethane repetitives of ester.In certain embodiments, the polymer is a kind of open-celled foam.
On the other hand, the present invention provides a kind of fit modified polymer or copolymer, the polymer or copolymer bagContain：
A) a kind of first repetitives, first repetitives have one or more hydroxyls；
B) a kind of second repetitives, second repetitives have one or more
-O-Si(R1)(R2)-R3- X- linkers-fit group, wherein：
R1And R2It is independently hydrogen, hydroxyl, halogen, alkyl(C≤6)Or alkoxyl(C≤6)；
R3It is alkane diyl(C≤20)；
X is-NH- or-S-；
The linker is-C (O)-alkane diyl(C≤12)-；And
It is a kind of protein of biological activity that this is fit, and the protein passes through amino, a hydroxyl or a mercaptoBase group is bound to the linker group.
In certain embodiments, first repetitives have two oh groups.
In certain embodiments, first repetitives have below formula：
In certain embodiments, R1And R2It is isopropyl.In certain embodiments, wherein R1And R2It is hydroxyl.In some realitiesIn applying example, R3It is alkane diyl(C3-11).In certain embodiments, R3It is-(CH2)3-。
In certain embodiments, second repetitives have below formula：
In certain embodiments, R3It is-(CH2)11-.In certain embodiments, second repetitives have following chemistryFormula：
In certain embodiments, X is-NH-.In certain embodiments, the linker is further defined by below formula：
Wherein carbonyl carbon is attached to X.
In certain embodiments, the fit mercapto groups by a cysteine residues are bound to the linker.In some embodiments, it is granulocyte-macrophage colony stimutaing factor (GM-CSF) that this is fit.
In certain embodiments, the fit modified polymer is further triple comprising the one kind the with below formulaMultiple unit：
Wherein X is：
- alkane diyl(C≤12)-；
- fragrant diyl(≤12)-；
- fragrant diyl(≤6)-CH2- fragrant diyl(≤6)-；Or
The substituted pattern of any these groups.
In certain embodiments, X is further defined as：
In certain embodiments, the third repeating unit is a polyurethane repetitives.In certain embodiments, thisThree repetitives are the polyurethane repetitives based on ether.In certain embodiments, the third repeating unit is a baseIn the polyurethane repetitives of ester.In certain embodiments, the polymer is a kind of open-celled foam.
On the other hand, the present invention provides a kind of wound insert, and the wound insert is included as above and below is retouchedA kind of fit modified polymer stated or copolymer.On the other hand, the present invention provides a kind of wound and applies part, and the wound applies partIncluding wound insert and a drop cloth of the skin for being configured to a wound for being attached to neighbouring patient.In some embodimentsIn, the wound applies part and further includes a fluid delivering pad, and the fluid delivering pad is configured to be attached to the drop cloth and oneFluid source, so that the fluid source may be actuated is delivered to a wound by a kind of fluid by the deposited part of the wound.
On the other hand, the present invention provides a kind of Wound treating apparatus, and the equipment includes that the wound applies part and is configured toThe fluid source that the wound applies part is attached to, so that the fluid source may be actuated and a kind of fluid is delivered to into the wound applyPart.In certain embodiments, the equipment further includes to be configured to be attached to the vacuum source that the wound applies part, so thatThe vacuum source may be actuated to apply negative pressure to the deposited part of the wound.In certain embodiments, the fluid is molten comprising a kind of featureLiquid, the solution strengthens target and combines or contribute to target being discharged into wound bed.In certain embodiments, the fluid includes lifeReason saline solution, the solution with slightly acidic pH, subalkaline pH, with different surfaces activating agent (for example, polysorbateEster), the solution of EDTA and/or EGTA.In certain embodiments, the fluid includes hypochlorous acid and hypochlorite ion.
A. define
As used in this, term " fit " refers not only to be bound to the oligonucleotide of certain target molecules or peptide molecule, andAlso refer to any part for being bound to a target molecule or ion.For example, as used in this, " fit " will be including EDTA (ethylenediaminesTetraacethyl), it is bound to Ca2+。
When using under the background in a chemical group, " hydrogen " means-H；" hydroxyl " means-OH；" oxo " mean=O；" halogen " independently means-F ,-Cl ,-Br or-I；" amino " means-NH2(referring to below for the base containing term aminoThe definition of group's (for example, alkyl amino))；" hydroxyl amino " means-NHOH；" nitro " means-NO2；Imino group means=NH (ginsengsSee the definition below for the group containing term imino group (for example, alkyl imino))；" cyano group " means-CN；" azido "Mean-N3；" phosphate " means-OP (O) (OH) under univalent background2Or the form of its deprotonation；" the phosphorus under bivalence backgroundHydrochlorate " means-OP (O) (OH) O- or the form of its deprotonation；" sulfydryl " means-SH；" sulfenyl " means=S；" thioether " anticipatesFinger-S-；" sulfonamido " means-NHS (O)2- (referring to below for (for example, the alkyl sulphur of the group containing term sulfonamidoAcylamino-) definition)；" sulfonyl " means-S (O)2- (referring to below for (for example, the alkyl of the group containing term sulfonylSulfonyl) definition)；And " sulfinyl " means-S (O)-(referring to below for the group (example containing term sulfinylSuch as, alkyl sulphinyl) definition).
Symbol "-" means a singly-bound, and "=" means a double bond, and " ≡ " means three keys.Symbol " ----" representOne optional key, the key is if it is present be singly-bound or double bond.SymbolRepresent a singly-bound or a double bond.Thus, for example, structureIncluding structure：AndSuch as the technology of this areaPersonnel will be understood that this annular atom of neither one forms a part for more than one double bond.When vertically drawing across a keyWhen, symbolIndicate an attachment point of the group.It should be noted that the attachment point typically only for more macoradical withThis mode is identified, to help reader rapid and clearly to identify an attachment point.SymbolMean oneSingly-bound, wherein being attached to the group " outside the page " of the butt end of the wedge.SymbolMean a singly-bound, wherein attachedIt is connected to the group " in the page " of the butt end of the wedge.SymbolMean a singly-bound, wherein conformation (for example, ROr S) or geometry is undefined (for example, E or Z).
Any undefined quantivalence on one atom of a shown in this application structure impliedly represents keyIt is bonded to a hydrogen atom of the atom.When a group " R " is depicted as one " floating group " in a loop systems, exampleSuch as, in below formula：
So R can substitute any hydrogen atom for being attached to any these annular atoms, including description, it is implicit,Or clearly defined hydrogen, as long as forming a stable structure.
When a group " R " is depicted as one " floating group " in the loop systems for condensing, such as withIn lower chemical formula：
Then unless otherwise indicated, otherwise R can substitute any these annular atoms for being attached to any one of these condensed ringAny hydrogen.Alternative hydrogen includes hydrogen (hydrogen of nitrogen for example, is attached in above chemical formula), the implicit hydrogen (example describedSuch as, it is not shown, it should be understood that the hydrogen of above the chemical formula for existing), clearly defined hydrogen and its exist depending on oneThe optional hydrogen (hydrogen of group X for example, being attached to, when X is equal to-CH-) of the identity of annular atom, if formed one it is stableStructure.In the illustrated case, R may reside within 5 yuan of the loop systems for condensing or 6 yuan of rings.Change more thanIn formula, the subscript letter " y " of the group " R " being followed by enclosed in bracket represents a numerical variable.Unless otherwise indicated,This variable can be 0,1,2 or the arbitrary integer more than 2, and the variable is only by the ring or the alternative hydrogen atom of loop systemsMaximum number limited.
For following these groups and classification, below further group/the class declaration is such as the subscript in bracketUnder：" (Cn) " defines the exact number (n) of the carbon atom in the group/classification." (C≤n) " is defined on can in the group/classificationThe maximum number (n) of the carbon atom to have, wherein for the group for being discussed, minimal amount is little as far as possible, for example, it should be appreciated thatGroup " thiazolinyl(C≤8)" or classification " alkene(C≤8)" in the minimal amount of carbon atom be two.For example, " alkoxyl(C≤10)" refer toWith from 1 to 10 carbon atom (for example, 1,2,3,4,5,6,7,8,9 or 10 or wherein can derivative any scope (exampleSuch as, 3 to 10 carbon atoms)) those alkoxy bases.(Cn-n') define the minimum (n) of carbon atom in the group with it is maximumNumber (n').Similarly, " alkyl(C2-10)" refer to from 2 to 10 carbon atoms (for example, 2,3,4,5,6,7,8,9 or 10,Or wherein can derivative any scope (for example, 3 to 10 carbon atoms)) those alkyl groups.
Except as indicated below, term " saturation " as used in this still means that the compound or base of such modificationGroup does not have carbon-to-carbon double bond and carbon-to-carbon triple bond.The term is not excluded for carbon-heteroatom multiple bond, such as one C=O bond orIndividual carbon-to-nitrogen double bon.Additionally, it is not excluded for being sent out possibly as the tautomeric part of ketoenol tautomerization or imines/enamineA raw carbon-to-carbon double bond.
When in no modifier, " when using in the case of " substituted ", term is " aliphatic " to represent the chemical combination so modifiedThing/group is an acyclic or ring-type but non-aromatic hydrocarbon compound or group.In aliphatic compound/group,Carbon atom can be bonded together by straight chain, side chain or non-aromatic ring (alicyclic).Aliphatic compound/group can beSaturation, i.e., (alkane/alkyl) is engaged by singly-bound；Or it is undersaturated, with one or more double bonds (alkene/thiazolinyl) or toolThere are one or more three keys (alkynes/alkynyl).When term " aliphatic " in the case of without modifier " substituted " when using,Only exist carbon and hydrogen atom.When the term and " substituted " modifier are used together, one or more hydrogen atoms are independentGround is by-OH ,-F ,-Cl ,-Br ,-I ,-NH2、-NO2、-CO2H、-CO2CH3、-CN、-SH、-OCH3、-OCH2CH3、-C(O)CH3、-N(CH3)2、-C(O)NH2Or-OC (O) CH3Substitute.
When using in the case of without modifier " substituted ", term " alkyl " is referred to a carbon atom as attachedContact, with straight chain or side chain, ring, ring-type or acyclic structure, and without in addition to carbon and hydrogenThe aliphatic group of the univalent saturation of of atom.Therefore, as used in this, cycloalkyl is a subset of alkyl.Group-CH3(Me)、-CH2CH3(Et)、-CH2CH2CH3(n- Pr) ,-CH (CH3)2(iso- Pr) ,-CH (CH2)2(cyclopropyl) ,-CH2CH2CH2CH3(n- Bu) ,-CH (CH3)CH2CH3(sec-butyl) ,-CH2CH(CH3)2(isobutyl group) ,-C (CH3)3(tertiary fourthBase) ,-CH2C(CH3)3(neopentyl), cyclobutyl, cyclopenta, cyclohexyl and cyclohexyl methyl are the unrestricted of alkyl groupProperty example.When using in the case of without modifier " substituted ", term " alkane diyl " is referred to one or two saturationCarbon atom as one or more attachment points, with straight chain or side chain, ring, ring-type or acyclic structure,Without carbon-to-carbon double bond or three keys, and the not aliphatic group of a bivalence saturation of the atom in addition to carbon and hydrogen.BaseGroup-CH2- (methylene) ,-CH2CH2-、-CH2C(CH3)2CH2-、-CH2CH2CH2- andIt is alkane diyl groupNon-limiting examples.When without using in the case of modifier " substituted ", term " alkylidene " refer to divalent group=CRR', wherein R and R' are independently that hydrogen, alkyl, or R and R' represent together with least two carbon atoms alkane diyl.The non-limiting examples of alkane diyl group include：=CH2,=CH (CH2CH3) and=C (CH3)2.When any these terms withWhen modifier " substituted " is used together, one or more hydrogen atoms are independently by-OH ,-F ,-Cl ,-Br ,-I ,-NH2、-NO2、-CO2H、-CO2CH3、-CN、-SH、-OCH3、-OCH2CH3、-C(O)CH3、-N(CH3)2、-C(O)NH2Or-OC (O) CH3ReplaceGeneration.Following group is the non-limiting examples of the alkyl group for replacing：-CH2OH、-CH2Cl、-CF3、-CH2CN、-CH2C(O)OH、-CH2C(O)OCH3、-CH2C(O)NH2、-CH2C(O)CH3、-CH2OCH3、-CH2OC(O)CH3、-CH2NH2、-CH2N(CH3)2、And-CH2CH2C1.Term " fluoroalkyl " is a subset of the alkyl for replacing, and wherein one or more hydrogen are by a fluorineBase replaces and in addition to carbon, hydrogen and fluorine, there are no other atoms.Group-CH2F、-CF3And-CH2CF3It is fluoroalkylThe non-limiting examples of group." alkane " refers to that compound H-R, wherein R are alkyl.
When using in the case of without modifier " substituted ", term " thiazolinyl " is referred to a carbon atom as attachedContact, with straight chain or side chain, ring, ring-type or acyclic structure, the carbon with least one non-aromatic-Carbon double bond, no carbon-to-carbon triple bond, and the not undersaturated aliphatic group of a unit price of the atom in addition to carbon and hydrogen.The non-limiting examples of alkenyl group include：- CH=CH2(vinyl) ,-CH=CHCH3,-CH=CHCH2CH3、-CH2CH=CH2(pi-allyl) ,-CH2CH=CHCH3And-CH=CH-C6H5.When using in the case of no modifier " substituted ",Term " alkene diyl " referred to two carbon atoms as attachment point, with straight chain or side chain, ring, ring-type or non-The structure of ring-type, the carbon-to-carbon double bond with least one non-aromatic, without carbon-to-carbon triple bond and not in addition to carbon and hydrogenAtom the undersaturated aliphatic group of a bivalence.Group-CH=CH- ,-CH=C (CH3)CH2- ,-CH=CHCH2-, withAndIt is the non-limiting examples of alkene diyl group.When these terms are used together with modifier " substituted "When, one or more hydrogen atoms are independently by-OH ,-F ,-Cl ,-Br ,-I ,-NH2、-NO2、-CO2H、-CO2CH3、-CN、-SH、-OCH3、-OCH2CH3、-C(O)CH3、-N(CH3)2、-C(O)NH2Or-OC (O) CH3Substitute.Group-CH=CHF ,-CH=CHCl and-CH=CHBr is the non-limiting examples of the alkenyl group for replacing." alkene " refers to that compound H-R, wherein R are alkeneBase.
Make with an aromatic carbon atom when when using in the case of without modifier " substituted ", term " aryl " is referred toFor the undersaturated aromatic group of a unit price of attachment point, the carbon atom forms one or more hexa-atomic aromatic ring structuresA part, wherein these annular atoms are all carbon, and the wherein group is not comprising the atom in addition to carbon and hydrogen.If there is manyIn a ring, then these rings can be condense or non-condensed.As used in this, the term is not excluded for being attached to first fragrantThe presence (carbon number is limited to be allowed) of one or more alkyl groups of fragrant ring or existing any other aromatic rings.Aryl baseThe non-limiting examples of group include phenyl (Ph), aminomethyl phenyl, (dimethyl) phenyl ,-C6H4CH2CH3(ethylphenyl), naphthyl,And the monoradical derived from xenyl.When using in the case of without modifier " substituted ", term " fragrant diyl " isRefer to two aromatic carbon atoms as attachment point an O divalent aromatic group, the carbon atom formed one or more sixA part for first aromatic ring structure, wherein these annular atoms are all carbon, and wherein the monoradical not comprising except carbon and hydrogen withOuter atom.As used in this, the term is not excluded for being attached to the first aromatic rings or existing any other aromatic ringsThe presence (carbon number is limited to be allowed) of one or more alkyl groups.If more than a ring, then these rings can be thickIt is closing or non-condensed.The non-limiting examples of fragrant diyl group include：
And
When these terms and modifier " substituted " are used together, one or more hydrogen atoms independently by-OH、-F、-Cl、-Br、-I、-NH2、-NO2、-CO2H、-CO2CH3、-CN、-SH、-OCH3、-OCH2CH3、-C(O)CH3、-N(CH3)2、-C(O)NH2Or-OC (O) CH3Substitute." aromatic hydrocarbons " refers to that compound H-R, wherein R are aryl.
When using in the case of without modifier " substituted ", term " aralkyl " refers to monoradical-alkane diyl-virtueBase, wherein term alkane diyl and aryl are individually to be used in the mode consistent with definition presented above.AralkylNon-limiting examples be：Benzyl (benzyl, Bn) and 2- phenyl-ethyl groups.When the term makes together with modifier " substituted "Used time, from alkane diyl and/or aryl one or more hydrogen atoms independently by-OH ,-F ,-Cl ,-Br ,-I ,-NH2、-NO2、-CO2H、-CO2CH3、-CN、-SH、-OCH3、-OCH2CH3、-C(O)CH3、-N(CH3)2、-C(O)NH2Or-OC (O)CH3Substitute.The non-limiting examples of substituted aralkyl are：(3- chlorphenyls)-methyl and the chloro- 2- phenyl -ethyl- 1- bases of 2-.
When using in the case of without modifier " substituted ", term " heteroaryl " is referred to an aromatic carbon atomOr nitrogen-atoms are used as a monovalent aromatic radical of attachment point, the carbon atom or nitrogen-atoms one aromatic ring structure of formationA part, wherein at least one of these annular atoms are nitrogen, oxygen or sulfur, and the wherein group is not comprising except carbon, hydrogen, fragranceAtom beyond nitrogen, fragrant oxygen and fragrant sulfur.As used in this, the term is not excluded for being attached to the aromatic rings or existingAny other aromatic rings one or more alkyl groups presence (carbon number limit allow).Heteroaryl groups it is unrestrictedProperty example include furyl, imidazole radicals, indyl, indazolyl (Im), picolyl, oxazolyls, pyridine radicals, pyrrole radicals, phoneticPiperidinyl, pyrazinyl, quinolyl, quinazolyl, quinoxalinyl, thienyl and triazine radical.When having modifier " substituted " in narrow eyes into a slitIn the case of when using, term " heteroaryl diyl " is referred to two aromatic carbon atoms, two fragrant nitrogen-atoms or an aromatic carbonUsed as an O divalent aromatic group of attachment point, the atom forms one or more fragrance to the fragrant nitrogen-atoms of atom andA part for ring structure, wherein at least one of these annular atoms are nitrogen, oxygen or sulfur, and wherein the divalent group does not includeAtom in addition to carbon, hydrogen, aromatic nitrogen, fragrant oxygen and fragrant sulfur.As used in this, the term is not excluded for being attached to firstThe presence (carbon number is limited to be allowed) of one or more alkyl groups of aromatic rings or existing any other aromatic rings.IfThere is more than one ring, then these rings can be condense or non-condensed.The non-limiting examples bag of heteroaryl diyl groupInclude：
And
When these terms and modifier " substituted " are used together, one or more hydrogen atoms independently by-OH、-F、-Cl、-Br、-I、-NH2、-NO2、-CO2H、-CO2CH3、-CN、-SH、-OCH3、-OCH2CH3、-C(O)CH3、-N(CH3)2、-C(O)NH2Or-OC (O) CH3Substitute.
When using in the case of without modifier " substituted ", term " acyl group " refers to that group-C (O) R, wherein R areHydrogen, alkyl, aryl, aralkyl or heteroaryl, these terms are as defined above.Group-CHO ,-C (O) CH3 (acetyl group,Ac)、-C(O)CH2CH3、-C(O)CH2CH2CH3、-C(O)CH(CH3)2、-C(O)CH(CH2)2、-C(O)C6H5、-C(O)C6H4CH3、-C(O)CH2C6H5,-C (O) (imidazole radicals) be carboxyl groups non-limiting examples." sulfonyl " is the side to be similar toFormula is defined, except the oxygen atom of group-C (O) R substitutes (- C (S) R) by a sulphur atom.When in these termsWhen any one is used together with modifier " substituted ", one or more hydrogen atoms independently by-OH ,-F ,-Cl ,-Br ,-I、-NH2、-NO2、-CO2H、-CO2CH3、-CN、-SH、-OCH3、-OCH2CH3、-C(O)CH3、-N(CH3)2、-C(O)NH2Or-OC(O)CH3Substitute.Group-C (O) CH2CF3、-CO2H (carboxyl) ,-CO2CH3(methyl carboxyl)-CO2CH2CH3、-C(O)NH2(ammonia firstAcyl group) and-CON (CH3)2It is the non-limiting examples of the carboxyl groups for replacing.
When using in the case of without modifier " substituted ", term " alkoxyl " refers to that group-OR, wherein R are oneIndividual alkyl, the term is as defined above.The non-limiting examples of alkoxy base include：-OCH3、-OCH2CH3、-OCH2CH2CH3、-OCH(CH3)2、-OCH(CH2)2,-O-ring amyl group and-O-ring hexyl.When without modifier " substituted "In the case of when using, term " alkenyloxy group ", " alkynyloxy group ", " aryloxy group ", " aralkoxy ", " heteroaryl epoxide " and " acyl-oxygenBase " refers to the group for being defined as-OR, and wherein R is correspondingly thiazolinyl, alkynyl, aryl, aralkyl, heteroaryl and acyl group.ClassAs, when when using in the case of without modifier " substituted ", term " alkylthio group " refers to that group-SR, wherein R are an alkaneBase, the term is as defined above.When using in the case of without modifier " substituted ", term " alkoxyl diyl " is referred toDivalent group-O- alkane diyls-.When any these terms are used together with modifier " substituted ", one or more hydrogen atomsIndependently by-OH ,-F ,-Cl ,-Br ,-I ,-NH2、-NO2、-CO2H、-CO2CH3、-CN、-SH、-OCH3、-OCH2CH3、-C(O)CH3、-N(CH3)2、-C(O)NH2Or-OC (O) CH3Substitute.Term " alcohol " corresponds to alkane as defined above, wherein extremelyA few hydrogen atom is substituted by an oh group.
When using in the case of without modifier " substituted ", term " alkyl amino " refers to that group-NHR, wherein R areOne alkyl, the term is as defined above.The non-limiting examples of alkylamino group include：-NHCH3With-NHCH2CH3。When using in the case of without modifier " substituted ", term " dialkyl amido " refers to group-NRR', and wherein R and R' canBeing same or different alkyl group, or R and R' can together represent an alkane diyl.The non-limit of dialkyl amino groupProperty example processed includes：-N(CH3)2、-N(CH3)(CH2CH3) and N- pyrrolidinyls.When in the feelings without modifier " substituted "When using under condition, term " alkoxy amino ", " alkenyl amino ", " alkynylamino ", " arylamino ", " aryl alkyl amino ", " miscellaneousArylamino " and " alkyl sulfonyl-amino " refer to the group for being defined as-NHR, wherein R be correspondingly alkoxyl, thiazolinyl,Alkynyl, aryl, aralkyl, heteroaryl and alkyl sulphonyl.The non-limiting examples of arylamino groups are-NHC6H5.WhenWhen using in the case of without modifier " substituted ", term " amide groups (amido) " (acyl amino (acylamino)) is referred toGroup-NHR, wherein R are acyl groups, and the term is as defined above.The non-limiting examples of amide group are-NHC (O) CH3。When using in the case of without modifier " substituted ", term " alkyl imino " refers to that divalent group=NR, wherein R areOne alkyl, the term is as defined above.When any these terms are used together with modifier " substituted ", one or manyIndividual hydrogen atom is independently by-OH ,-F ,-Cl ,-Br ,-I ,-NH2、-NO2、-CO2H、-CO2CH3、-CN、-SH、-OCH3、-OCH2CH3、-C(O)CH3、-N(CH3)2、-C(O)NH2Or-OC (O) CH3Substitute.Group-NHC (O) OCH3With-NHC (O) NHCH3It is the non-limiting examples of the amide group for replacing.
When without using in the case of modifier " substituted ", term " alkyl sulphonyl " and " alkyl sulphinyl " phaseRefer to group-S (O) with answering2R and-S (O) R, wherein R is an alkyl, and the term is as defined above.Term " thiazolinyl sulphonylBase ", " alkynylsulfonyl ", " aryl sulfonyl ", " arylalkyl sulfonyl " and " heteroarylsulfonyl " are in a similar mannerIt is defined.When any these terms are used together with modifier " substituted ", one or more hydrogen atoms are independentlyBy-OH ,-F ,-Cl ,-Br ,-I ,-NH2、-NO2、-CO2H、-CO2CH3、-CN、-SH、-OCH3、-OCH2CH3、-C(O)CH3、-N(CH3)2、-C(O)NH2Or-OC (O) CH3Substitute.
Term " glucosides " refers to that one of glycosyl is bound to a kind of compound of a non-carbohydrate part.Typical caseGround, the glycosyl (glycone) is to be bonded to another group by its anomeric carbon via a glycosidic bond (glucosides is matched somebody with somebodyBase), the glycosidic bond has oxygen, nitrogen or a sulphur atom as linker.
" simple sugars (simple sugar) " is the basic structural unit of carbohydrate, and these units can not easily waterSolution is into simpler unit.A kind of basic chemical formula of simple monosaccharide is CnH2nOn, wherein Integer n is at least 3 and seldom bigIn 7.Simple monosaccharide can generically be named according to the number of carbon atom n：Triose, tetrose, pentose, hexose, etc..LetterMonosaccharide can be open chain (acyclic), ring-type or its mixture.In these annular forms, the ring generally has 5 or 6Individual atom.These forms are correspondingly referred to as furanose and pyranose --- by carrying out analogy with furan and pyrans.Simple sugars canTo be categorized further, as aldose class (have in the end of the chain of non-annularity form carbonyl group those) and ketose class(wherein carbonyl group not the end of chain those).The non-limiting examples of aldose include：Glycolaldehyde, glyceraldehyde, red mossSugar, threose, ribose, arabinose, xylose, lyxose, allose, altrose, glucose, mannose, gulose, idose,Galactose and talose.The non-limiting examples of ketose include：Dihydroxy acetone, Erythrulose, ribulose, xylulose, fruitSugar, psicose, sorbose and Tagatose.' D- ' and ' L- ' prefix can be used for distinguishing be mutual mirror image two kinds of spiesFixed stereoisomer.Term simple sugars are also contemplated by its O- acetyl derivative.
" amino sugar " or " aminoglycoside " refers to a kind of derivant of a kind of sugar, deoxysaccharide, saccharic acid or sugar alcohol, wherein oneIndividual or multiple oh groups are substituted by one or more amino groups." simple amino sugar " refers to a kind of simple sugars, simpleA kind of derivant of deoxysaccharide, simple saccharic acid or sugar alcohol, wherein one or more oh groups are by one or more aminoGroup is substituted.These terms are also contemplated by itself N- and O- acetyl derivative.Non-limiting examples include N- acetyl glucosamines, halfLactose amine, glycosamine and sialic acid.
Term " deoxysaccharide " refers to a kind of sugar derivativess, and in the oh group of one of which carbohydrate isSubstituted by a hydrogen atom." simple deoxysaccharide " is a kind of deoxysaccharide derived from a kind of simple sugars as defined in this.ThisA little terms are also contemplated by its O- acetyl derivative.The non-limiting examples of simple deoxysaccharide are deoxyribose (based on ribose), seaAlgae sugar and rhamnose.
Term " saccharic acid " refers to a kind of sugar derivativess, and one of aldehyde functional group or one or more hydroxy functional groups areIt is oxidized to a carboxylic group.Aldose acid is oxidized those of aldehyde functional group of wherein aldose.Onosic acid is whereinFirst oh group of 2- ketoses is oxidized, so as to produce a kind of 2-ketoacid those.Alduronic acid is wherein aldose or ketoneOxidized those of the terminal hydroxyl of candy.Aldaric acid is two ends all oxidized those of wherein aldose.Non- limitProperty glycuronic acid processed includes glyceric acid (3C), xylonic (5C), gluconic acid (6C) and ascorbic acid (6C, unsaturated lactone).KetoneThe non-limiting examples of saccharic acid include neuraminic acid (5- amino -3,5- dideoxy-D- glyceryls-D- galas-nonyl- 2- onosic acid)With ketone group deoxidation ketooctulosonic acid (KDO or 3- deoxidations-D- manna-octyl- 2- onosic acid).The non-limiting examples of alduronic acid include PortugalAlduronic acid (6C), galacturonic acid (6C) and iduronic acid (6C).The non-limiting examples of aldaric acid include tartaric acid(4C), meso-galactosaccharic acid (galactosaccharic acid) (6C) and D- glucosaccharic acids (glucosaccharic acid) (6C)." simple sugarsAcid " is a kind of saccharic acid derived from a kind of simple sugars.These terms are also contemplated by its O- acetyl derivative.
Term " sugar alcohol " refers to a kind of sugar derivativess, the carbonyl group (aldehydes or ketones, reducing sugar) of the sugar derivativess byIt is reduced into a primary or secondary oh group.The non-limiting examples of sugar alcohol include：It is ethylene glycol (2 carbon), glycerol (3 carbon), redIt is moss sugar alcohol (4 carbon), threitol (4 carbon), 1,2,3,4,5-pentanepentol (5 carbon), xylitol (5 carbon), ribitol (5 carbon), sweetDew sugar alcohol (6 carbon), Sorbitol (6 carbon), dulcitol (6 carbon), iditol (6 carbon), hydroxyl isomaltulose (12Carbon), maltose alcohol (12 carbon), lactose (12 carbon) or hydrogenated glucose (polyglycitol)." simple sugar alcohol " is to spread outIt is conigenous a kind of a kind of sugar alcohol of simple sugars.These terms are also contemplated by its O- acetyl derivative.
As used in this, term " monosaccharide group " refers to a univalent carbon aquation with a carbon atom as attachment pointCompound group.The term covers the group produced from the removal of following thing by an oh group：A kind of simple sugars (exampleSuch as, glucose), simple deoxysaccharide (for example, trehalose), simple saccharic acid (for example, gluconic acid), simple sugar alcohol (for example, xyloseAlcohol) or simple amino sugar (for example, glycosamine).Typically, the anomeric carbon that the monosaccharide group passes through it via oxygen atom linkerIt is bonded to another group (aglycone).In some cases, the linker can be a nitrogen or sulphur atom.
The univalent carbohydrate group that " disaccharide group " is made up of two monosaccharide groups, wherein the second monosaccharide groupsGroup substitutes an oxygen on an oh group of the first monosaccharide group.The non-limiting examples of disaccharide group are included derived from sugarcaneThose of sugar, lactulose, Lactose, maltose, trehalose and cellobiose.
The univalent carbohydrate group that " trisaccharide group " is made up of three monosaccharide groups, wherein the second monosaccharide groupsGroup substitutes a hydrogen on an oh group of the first monosaccharide group, and the 3rd monosaccharide group substitute this first or thisA hydrogen on one oh group of two monosaccharide groups.
The univalent carbohydrate-based that oligosaccharide is made up of three to ten, preferably three to six monosaccharide groupsGroup, wherein the second monosaccharide substitute the first monosaccharide an oh group on a hydrogen, the 3rd monosaccharide substitute this first or shouldA hydrogen on one oh group of the second monosaccharide group, and the monosaccharide groups engaged before follow-up monosaccharide group is substituted arbitrarilyHydrogen in group, therefore form a kind of straight chain or side chain structure.
When using in the case of without modifier " substituted ", term " silicyl " refers to group-SiR3, wherein oftenIndividual R is independently hydrogen or unsubstituted alkyl, and the term is as defined above.Term " substituted silicyl " refer to group-SiR3, wherein at least one of these R groups and these R groups is up to independently all the alkyl of a replacementOr-OH ,-F ,-Cl ,-Br ,-I ,-NH2、-NO2、-CO2H、-CO2CH3、-CN、-SH、OCH3、-OCH2CH3、-C(O)CH3、-N(CH3)2、-C(O)NH2Or-OC (O) CH3.Any remaining R group of the substituted silyl-group is independently hydrogen or notSubstituted alkyl.Term " silylated " or " silanization " indicate that a kind of given compound is derivatized into and includeOne silicyl and/or substituted silyl-group.Abbreviation "-Sil " refers to silicyl and/or substituted silicylGroup.
Term " coupled " is defined as being concatenated, but might not directly, and not necessarily mechanically；Two objects of " connection " are that can be in one individual.Term " one (a) " and " a kind of (an) " are defined as one or more,Unless this disclosure is clearly required otherwise.Term " substantially ", " about " and " about " is defined as to a great extent but nothingSpecified by need to complying fully with, this will be understood by those skilled in the art.
Term " including (comprise) " (and any form for including, such as " containing (comprises) " and " include(comprising) ", " have (have) " (and any form having such as " being provided with (has) " and " has(having) "), " including (include) " (and including any form, such as " including (includes) " and " including") and " contain (contain) " (and any form for containing such as " containing (contains) " and " contains (including)Have (containing) ") it is open copulative verb.Therefore, a kind of "comprising", " having ", " including " or " containing " oneThe method of individual or multiple steps possesses the one or more step, but is not limited to only possess the one or more step.Similarly,A kind of "comprising", " having ", the deposited part of the wound of " including " or " containing " one or more elements possess the one or more unitPart, but be not limited to only possess these elements.For example, part is applied in a kind of wound including a kind of wound insert and a drop clothIn, the wound applies the element that part specified including these but is not limited to only have these elements.For example, such wound applies part alsoA connection gasket can be included, the connection gasket is configured to be attached on a Wound treating apparatus.
As term " effective " is used in this specification and/or claims, the term means to be enough to realize one kindDesired, expected or desired result.
As used in this, term " IC50" a kind of inhibition dosage is referred to, the dosage is obtained maximum reaction50%.This quantitative measurement indicate by a given biological, biochemistry or chemical process (or the component of a process, i.e. enzyme,Cell, cell receptor or microorganism) suppress a kind of concrete medicine or other materials (inhibitor) required for halving to measure.
As used in this, term " patient " or " experimenter " refer to a mammalian organism living, such as people, monkey, cattle,Sheep, goat, Canis familiaris L., cat, mice, rat, Cavia porcelluss or its genetically modified organism.In certain embodiments, the patient or experimenter are onePlant primate.The non-limiting examples of people experimenter are adult, teenager, baby and fetus.
" PBS " is phosphate buffered saline (PBS)；" phr " number parts per hundred parts in resin；" OPA " is o-phthalaldehyde(OPA)(ortho-pthaldialdehyde)；" RT " is room temperature.
" pharmaceutically acceptable " that here is such as generally used refer to be suitable in rational medical judgment scope withThe tissue of human and animal, organ, and/or body fluid contact and use, and without excessive toxicity, stimulation, anaphylaxiss or other askTopic or complication, while those compounds with the reasonable benefit/Hazard ratio for matching, material, compositionss, and/or dosage form.
" pharmaceutically acceptable salt " means pharmaceutically acceptable as defined above and possesses desiredThe salt of the compound of the invention of pharmacologically active.This kind of salt is included with mineral acid (such as hydrochloric acid, hydrobromic acid, sulphuric acid, nitric acid, phosphoric acidDeng) formed acid-addition salts；Or the acid-addition salts formed with following organic acid：Such as 1,2- ethionic acids, 2- ethylenehydrinsulfonic acids, 2-LOMAR PWA EINECS 246-676-2,3- phenylpropionic acids, 4,4' methylene bis (3- hydroxyl -2- alkene -1- formic acid), 4- methyl bicycles [2.2.2] oct-2-ene -L- formic acid, acetic acid, aliphatic list and dicarboxylic acids, aliphatic sulphuric acid, elixir of vitriol, benzenesulfonic acid, benzoic acid, camphorsulfonic acid, carbonAcid, cinnamic acid, citric acid, Pentamethylene. propanoic acid, ethyl sulfonic acid, fumaric acid, glucoheptonic acid, gluconic acid, glutamic acid, glycolic, enanthic acid,It is caproic acid, hydroxynaphthoic acid, lactic acid, lauryl sulfate, maleic acid, malic acid, malonic acid, mandelic acid, methanesulfonic acid, muconic acid, o-(4- hydroxy benzoyls) benzoic acid, oxalic acid, p- chlorobenzenesulfonic acid, phenyl replace alkanoic acid, propanoic acid, p- toluenesulfonic acid, thirdKeto acid, salicylic acid, stearic acid, succinic acid, tartaric acid, butylacetic acid, trimethylace tonitric etc..Pharmaceutically acceptable salt is also wrappedInclude base addition salts, these base addition salts can existing sour proton can with inorganic or organic alkali reaction when formed.Can connectThe inorganic base received includes sodium hydroxide, sodium carbonate, potassium hydroxide, aluminium hydroxide and calcium hydroxide.Acceptable organic base bagInclude ethanolamine, diethanolamine, triethanolamine, trometamol, N-METHYL-ALPHA-L-GLUCOSAMINE etc..It will be appreciated that forming any of the present inventionThe concrete anion of a part for salt or cation are not crucial, as long as the salt is pharmacologically acceptable as an entirety.The other example of pharmaceutically acceptable salt and their preparation method and purposes are presented on pharmaceutical salts handbook：CharacteristicWith purposes (Handbook of Pharmaceutical Salts:Properties, and Use) (PH Si tal fibres(P.H.Stahl) edit with CG Wei Muts (C.G.Wermuth), Verlag Helvetica Chimica Acta are publishedSociety, 2002) in.
" prevention (Prevention) " or " prevention (Preventing) " includes：(1) suppress a kind of disease tested at oneOutbreak in person or patient, the experimenter or patient may be risky and/or be susceptible to suffer from the disease, should but also do not experience or showWhole pathology of disease or symptom, and/or (2) slow down the pathology or symptom of a kind of disease in an experimenter or patient send outWork, the experimenter or patient may be risky and/or be susceptible to suffer from the disease, but also do not experience or show whole pathology of the diseaseOr symptom.
As used in this, term " polymer " " includes " copolymer ".
" repetitives " are the simplest structural solids of some materials, the material such as framework and/or polymer, nothingBy being organic, inorganic or metal-organically.In the case of polymer chain, repetitives are continuously connected to along the chainTogether, just as the beadlet of necklace.For example, in polyethylene-[CH2CH2]n- in, repetitives are-CH2CH2-.Subscript " n " refers to poly-It is right, i.e. the number of the repetitives for linking together.When the value of " n " is undefined or wherein " n " is not present, it is simplyRefer to repetition and the aggregation property of material of chemical formula in bracket.The concept of repetitives is equally applicable to situations below,Connectivity three-dimensional extension wherein between these repetitives, such as in metal organic frame, modified polymer, thermosetting polymerizationIn thing etc..
Term " reticulated, open type foam " or ROCF refer to many with the one kind being made up of the interconnecting webs of solid strutsA kind of foamed materialss of pore structure.Perforate is formed by network structure chemical industry skill, and these perforates are further defined as placeIn a kind of form of net or with a kind of net being made up of some.Because all pillars are to link,Open-cell porosity is also what is linked, so as to produce a kind of continuously porous material.ROCF can be concrete by three independent characteristicsGround definition：Pore size, relative density and matrix material.In certain embodiments, ROCF be by polyurethane into.
The definition of any conflict in any reference that replacement defined above is incorporated herein by reference.However, defining certainThe fact that a little terms, is not construed as indicating that undefined any term is indefinite.But, all terms for being used are allIt is considered as to describe the present invention in the way of term so that one of ordinary skill in the art is it will be appreciated that the scope of the present inventionAnd put into practice the present invention.
In addition, the device for configuring in some way or structure are at least configured by that way, but it also may be usedTo be configured by the other modes in addition to those being specifically described.
B. fit modified material
In one aspect, the present invention provides a kind of covalent system for catching and concentrating the specificity factor in wound climateSystem, these factors include causing a kind of factor of notable biological respinse (such as propagation, differentiation or angiogenesis).This technology canBiomolecule or bioactive compound are delivered to into wound bed for (such as).Provided herein fit modified material canIt is a kind of for catching and concentrating a kind of (for example, the biology of initiation of the specificity factor in wound climate to be used as in certain embodimentsThe factor of reaction such as angiogenesis) or endogenouss or exogenous biomolecule or bioactive compound are delivered to into the woundThe system of bed.
In one aspect, there is provided fit modified material, these materials include at least three kinds components：(1) a kind of polymer,(2) at least one linker of the polymer is covalently attached to, for example, directly through a side chain or logical of the polymerAnother linker molecule for being linked to the polymer, and (3) at least one aptamer molecules are crossed, the molecule can covalently be attachedTo the main polymer chain, but more typically will covalently be attached to a linker.
1) suitable polymer
For the polymer that is used in conjunction with include hydrophobicity or hydrophilic polyurethane, shitosan, crosslinking and/Or uncrosslinked polyolefin, polyhydric alcohol, ethylene vinyl acetate (EVA), it is elastomer such as acrylonitrile butadiene rubber (NBR), poly-Chlorobutadiene (PCP or CR), ethylene propylene rubber (EPR and EPDM), poloxamer, silicone, and/or fluorocarbon polymer.For example,In certain embodiments, it is possible to use a kind of polyether polyols based on shitosan.
Polyurethane is reaction polymer.One ammonia ester bond is by making an isocyanate groups-N=C=O and a hydroxylBase radical reaction producing, and polyurethane be by a kind of polyisocyanate with a kind of dihydroxylic alcohols or a kind of polyhydric alcohol typicallyA kind of polyaddition reaction in the case where there is catalyst and other additives is producing.Polyisocyanate is that have twoOr a kind of molecule R- (N=C=O) of more isocyanate functional groupsn, wherein n >=2 and polyhydric alcohol is that have two or moreA kind of molecule R'- (OH) of multiple hydroxy functional groupsn, wherein >=2.The product is containing ammonia ester bond (- RNHCOOR'-)A kind of polymer.Polyurethane can be by making the one of a kind of liquid isocyanate and polyhydric alcohol, catalyst and other additivesPlant liquid blend to be reacted to produce.Polyhydric alcohol can also be referred to as a kind of resin or one kind with the blend of other additivesResin blend.In certain embodiments, resin blend additive can comprising chain elongation agent, cross-linking agent, surfactant,Fire retardant, foaming agent, pigment, and/or filler.Breathable (breathable) or breathability (air-permeable) are openedThe synthesis of pass flexibility urethane polymers is for example taught by United States Patent (USP) 5,686,501, and the patent is by quoting with itCombine here in full.
Molecule comprising two isocyanate groups is referred to as diisocyanate.Isocyanates can be categorized as aromatic series, such as methyl diphenylene diisocyanate (MDI), diphenylethane diisocyanate (EDI) or toluene di-isocyanate(TDI)(TDI)；Or it is aliphatic, such as the own diester of two Carbimide .s (HDI) or isophorone diisocyanate (IPDI).Aggretion type Carbimide.One example of ester is aggretion type methyl diphenylene diisocyanate, and it is with two, three and four or moreThe blend of the molecule of isocyanate groups.Isocyanates can pass through them partly with a kind of polyol reaction so as to shapeFurther it is modified into a kind of prepolymer." quasi-prepolymer " is to be more than 2 with the stoichiometric proportion of oh group in isocyanates:1When formed." actual prepolymer " is to be equal to 2 in the stoichiometric proportion:Formed when 1.The key character of isocyanates includes themMolecular backbone, NCO content %, degree of functionality and viscosity.
Molecule comprising two oh groups is referred to as dihydroxylic alcohols.Example includes ethylene glycol (EG), 1,4- butanediols(BDO), diethylene glycol (DEG).Molecule comprising three oh groups is referred to as trihydroxylic alcohol.Example includes glycerol.Polyhydric alcohol sheetBody can be polymer.For example, they can contain hydroxyl or amino by expoxy propane (PO), oxirane (EO) at oneInitiator on base catalysiss addition or by a kind of binary acid (such as adipic acid) and glycol (such as ethylene glycol or dipropylene glycol(DPG) polyesterification reaction) is forming.The polyhydric alcohol extended with PO or EO is typically referred to as polyether polyol.By polyesterThe polyhydric alcohol for changing reaction formation is typically referred to as PEPA.The choosing of the molecular weight of initiator, prolongation agent and polyhydric alcoholSelect by typically affect it physical state and resulting polyurethane physical characteristics.The key character of polyhydric alcohol is themMolecular backbone, initiator, molecular weight, primary hydroxy group %, degree of functionality and viscosity.
One attribute of polyurethane is the ability that it can be transformed into foam.In certain embodiments, can also addA small amount of water is reaching this point.Water will be reacted to produce carbon dioxide with isocyanates, and the gas is filled and madeHole expansion produced in the mixed process.Can also use foaming agent, including some halogenated hydrocarbons such as HFC-245fa (1,1,1,3,3- pentafluoropropanes) and HFC-134a (1,1,1,2- tetrafluoroethane) and hydro carbons such as pentane.In certain embodiments, can be withChange feature of the polymer in foaming process using surfactant.
Although the characteristic of polyurethane is typically mainly determined by the selection of polyhydric alcohol that diisocyanate is producedAffect.For example, cure rate is generally by by the shadow of the reactivity of a given functional group and the number of functional isocyanate groupsRing.
When the line style difunctionality Polyethylene Glycol segment using commonly known as polyether polyol is to produce ammonia ester bond, typical caseGround produces softer, elasticity and more flexible polyurethane.If using multi-functional polyol, then typically produce moreFirm product, because these polyhydric alcohol produce a kind of three-dimensional cross-linked structure, this structure may be at again a kind of low closeThe form of degree foam.Control viscoelastic property (for example, by changing used catalyst and polyhydric alcohol) can produce memoryFoam, the memory foam is in skin temperature than more soft at room temperature.
In some embodiments of the invention, polyurethane foam is by isocyanates and polyhydric alcohol (typically toluene twoIsocyanates and a kind of multi-arm polyether polyol) polyreaction being formed.Can be to these component indexings(indexed), so that the isocyanate groups and oh group are in one to one ratio.
Shitosan is derived from ectoskeletal a kind of linear polysaccharide of Crustacean, it in certain embodiments can by withLower chemical formula is representing：
In certain embodiments, shitosan is to press (deacetylated list by the D- glucose of β-(the l-4)-connection of random distributionUnit) a kind of linear polysaccharide for constituting of and N- acetyl group-GLUCOSAMINE (acetylated unit).The presence of oh group and amino groupAllow to use diversified conjugated chemistry.For example, shitosan can be with diisocyanate or triisocyanate molecule (including twoIsocyanates or tri-isocyanate prepolymers) polymerization and/or it is copolymerized into a kind of polyether polyols or copolymer.
In certain embodiments, in addition to shitosan, it is possible to use different from a kind of polyhydric alcohol of shitosan.Shitosan is addedAdd to and produce ammonia ester bond between a kind of oh group of polyol resin on isocyanates and shitosan ring.Resulting foamObtainable shitosan segment on the pillar of the foam is included in, and gives the foam with some characteristics of shitosan.Additionally,Addition shitosan produces a kind of foam of excessive indexing (over-indexed foam), and the foam has in shitosan chainSuperfluous oh group and amine groups in section, that be available for other surface chemistry.For example, Fig. 7 is depicted based on TDI, shellPolysaccharide and VoranolTMThe repetitives of the polyurethane of the polyreaction of 3010 trihydroxylic alcohols.In certain embodiments, shell gathersSugar or another kind of dihydroxylic alcohols or polyhydric alcohol can be by a kind of diisocyanate molecules of every 1.0g or prepolymers 0.05 to 0.5g shellsThe ratio polymerization generating polyurethane of polysaccharide.The other details of this class method and example are provided among examples section below.
5,10,20 and 30PHR (per the grams of the shitosan of hectogram polyurethane resin) are produced and have testedPreparation.They are entered by being available for and the quantity of surface amine that is bonded conjugated with o-phthalaldehyde(OPA) by the foamRow is characterized.It has been found that Relative fluorescence units RFU is that obtainable primary amine is proportional on surface to the foam.
2) the modified main polymer chain of linker
In certain embodiments, the present invention provides the silyl-modified polymer for replacing.Example is included comprising being based onThose of the repetitives of following thing：Polyurethane (can be hydrophobic or hydrophilic), shitosan, crosslinking and/or notThe polyolefin of crosslinking, polyhydric alcohol, ethylene vinyl acetate (EVA), elastomer such as acrylonitrile butadiene rubber (NBR), polychlorostyrene fourthAlkene (PCP or CR), ethylene propylene rubber (EPR and EPDM), silicone, and/or fluorocarbon polymer.For example, in certain embodiments,A kind of polyether polyols or copolymer based on shitosan can be used.In certain embodiments, substituted silicyl baseGroup is to be attached to the polymer or copolymer by an oxygen atom.In other embodiments, substituted silyl-group is straightGround connection is attached to a carbon atom of the polymer or copolymer.
In certain embodiments, silyl-group (for example, (H that offer of the invention replaces2N(CH2)3)(i-Pr)2Si- and (H2N(CH2)11)(EtO)2Si-) modified polymeric material.In certain embodiments, material of the invention includes onePlant foam of polymers.The example of foamed materialss can include that the foam can be hydrophobic or parent based on the foam of polyurethaneAqueouss, including for example a kind of polyurethane foamed material based on shitosan.Other foams that may be adapted to include shitosan,Crosslinking and/or uncrosslinked polyolefin, polyhydric alcohol, ethylene vinyl acetate (EVA) and elastomer such as acrylonitrile butadieneRubber (NBR), polychloroprene (PCP or CR), ethylene propylene rubber (EPR and EPDM), silicone and fluorocarbon polymer.For example,In certain embodiments, it is possible to use a kind of polyurethane foam based on shitosan.In certain embodiments, substituted monosilaneBase group is to be attached to the foam by an oxygen atom.In other embodiments, substituted silyl-group is directly attachedIt is connected to a carbon atom of the foam.
These silylated polyurethane foams disclosed by here can be by the way that the one kind comprising this kind of group be gatheredOh group silylanizing on compound is being prepared.As used in this, silylanizing is by the first silicon of a replacementAlkyl group (R3Si-) be incorporated into a molecule among.It is related to be substituted on the compound with the silyl-group of a replacementA hydrogen (for example, the hydrogen of an oh group), for example, compound 3- aminopropyl diisopropyls Ethoxysilane will be with hydroxylBase group reacts under proper condition, to form a new covalent Si-O key, so as to by H2N(CH2)3Si(iPr)2- group connectsAn oxygen is connected to, the oxygen is further attached to the attachment point on molecule, and the attachment point was wherein once attached to before oh group.Do not receiveThe constraint of mechanism, the oxygen atom of the product can be the same oxygen atom of the oh group reactant.Referring to " (how I applyMy silane(How do I apply my Silane)) " cover this special catalogue (Gelest Catalog.) the 2006, the 19th toPage 20, they combine in full here by quoting with it.
How example 1 below can be deposited on 30PHR shitosans by the silyl-group that amino replaces if being explainedOn foam.Example 2 is illustrated how can be conjugated with the execution maleimide-mercaptan of the sulfo group-EMCS with below formula：
In certain embodiments, the N- hydroxysuccinimide esters (NHS esters) on an end of the sulfo group-EMCS moleculesCan be reacted with the free amino group group of the linker or polymer.Dimaleoyl imino on another end of the molecule- SH the radical reactions that group can be used on fit with for example a kind of peptide, to form stable thioether bond.The sulphur by this wayBase-EMCS can be used for for the peptide being connected to a kind of polymer or copolymer, including a kind of foam substrate.
Additionally, these peptides used in maleimide-mercaptan is conjugated typically have especially modified C lastEnd.For example, these C-terminal can use the Cys residues " end-blocking " with reactivity-SH groups to promote this conjugated.Sulfo group-In EMCS conjugatess, be typically excessively used sulfo group-EMCS, for example, with relative to a kind of organosilane cross-linking agent 50 to 100 ×Molar excess, to promote the reaction.This kind of reaction can be carried out for example under pH 7.0 to 7.5 and room temperature.This kind of method can be withFurther changed using the vitochemical principle and technology such as applied by those skilled in the art and optimized.ThisClass principle and technology are for example taught the Advanced Organic Chemistry in agate strange (March)：Reaction, mechanism and structure (AdvancedOrganic Chemistry:Reactions, Mechanisms, and Structure) in (2007), the document is also by drawingHere is combined in full to it.It is, for example possible to use other suitable amino-sulfenyl linkers.These linkers include, for exampleSM (PEG) 24, SM (PEG) 12, SM (PEG) 8, SM (PEG) 6, SM (PEG) 4, sulfo group-LC-SMPT, SM (PEG) 2, sulfo group-KMUS, LC-SMCC, LC-SPDP, sulfo group-LC-SPDP, SMPH, sulfo group-SMPB, SMPB, SMPT, SIAB, sulfo group-SIAB,EMCS, sulfo group-SMCC, SMCC, MBS, GMBS, sulfo group-GMBS, sulfo group-MBS, SPDP, SBAP, BMPS, AMAS and SIA.
In some embodiments of the invention, the silylated polymer by with other linker molecule (for example,Oligopeptide, oligomer) further it is modified.In certain embodiments, these other linkers are covalently attached to the monosilaneThe polymer of base or the main chain of copolymer.In other embodiments, the linker is covalently attached to the polymer or copolymerA side chain or side base on.In certain embodiments, the other linker is via the silylated polymer or commonOne functional group (for example a, amino group) attachment of the silyl-group of one replacement of polymers, so as to form one kindMaterial-linker1- linker2Modified material, the wherein linker1It is that a silyl-group for replacing is (also referred to as differentDifunctional silane cross-linking agent), as discussed in this those, and linker2It can be another dibasic monosilaneBase group or another type of linker molecule, such as oligopeptide.In certain embodiments, linker2Can be by linker1OnOne terminal amino group or a mercapto groups are anti-with EMCS (N- [ε-dimaleoyl imino hexylyloxy] succimide ester)Should be being formed.Referring to example 2 below.
The further functionalization of the boundary layer produced by (such as) silylanizing can be by by the silicyl baseGroup or linker2The other end of group be connected to it is fit, including as discussed further below " capture peptide ".
3) the modified main polymer chain of fit-linker
In certain embodiments, can be by conjugated to a kind of fit (including based on the fit of peptide) polymer or copolymer.Peptide is fit, and here is also known as " capture peptide ", can be used for protein target being combined in wound climate, and does not suppress themSignal transduction ability or other functions.This kind of capture peptide can be also used for capturing and increasing through the wound bed of a patientThe concentration of biological activity protein, for example, to strengthen or activating a kind of biological respinse of institute's targeting.These and other purposes existIllustrating in greater detail below.
In certain embodiments, by a linker be further linked to one or more it is different types of fit, so as toForm a kind of main polymer chain-linker-fit conjugatess.In certain embodiments, the linker is covalently attached to the polymerizationThe main chain of thing.In other embodiments, the linker is covalently attached in a side base of the polymer.In some embodimentsIn, by the fit main chain for being attached directly to the polymer, so as to form a kind of main polymer chain-fit conjugatess.
Poloxamer-EMCS- peptidic constructs are also successfully used to be bound to a kind of foam by fit.Poloxamer it is hydrophobicPart has strong hydrophobic interaction with foam.Hydrophilic " arm " has amino group, then will using an EMCS linkerThese amino groups are connected to oligopeptide.
In certain embodiments, it is a kind of by calculating " capture peptide " derived from selection or phage display that this is fit.This kind of " capture peptide " can be used for targeting for combine specific protein or protein familieses.One example of this peptide isBy aminoacid sequence GHQGQHIGQMS-AEEAc-Cys-NH2Defined P-16, wherein AEEAc-Cys-NH2It is for conjugatedTo an especially modified C-terminal of the maleimide base group provided by the EMCS linkers.Have shown that P-16 is tiedIt is bonded to VEGF and PDGF and potentially other members of the PDGF superfamilies.
A kind of fit and 30PHR shitosans foam of example 2 below report is via a kind of sulfo group-EMCS ([N- ε-maleimideAmido hexylyloxy] sulfo group succimide ester) cross-linking agent it is conjugated.
For example, once with reference to, the 3- aminopropyls-diisopropyl Ethoxysilane terminal amino group keep can and,For capturing peptide or fit combination.In certain embodiments, biologically active peptide can by retain their biological activity oneThe mode of kind combines so far terminal amido group.When the material is used as a part for for example a kind of deposited part, a kind of wound insert or padWhen, the peptide of these attachments can be used for capturing and increasing the concentration of the biological activity protein through a wound bed, so as to increaseStrong or activation institute targeting biological respinse.For example, deposited part can be prepared for protein such as VEGF collectionIn in wound bed, so as to cause angiogenesis and/or cause micro-pipe formed.By this way, this kind of material can be used for moreThe different biological approaches in a wound climate are controlled well
When a kind of peptide or a kind of factor are attached to the other end of the silane crosslinker, it will be with one group of specified criteriaUnder the target molecule specific dissociation constant (K given tod) or binding affinity.In certain embodiments, it may be allowedThe Reversible binding of one or more target.In certain embodiments, when as the one of this these method and apparatus coveredDuring part, above-mentioned target can be discharged back into a wound.In certain embodiments, Multiple components can be added to instillationSolution, not only to help that the factor for being combined dissociation is back in the wound climate, but also is retained with one or moreExudate key element cooperative interaction for a kind of regulating and controlling effect so that drawing a kind of favourable wound healing reaction.The type of instillation solution is discussed in further detail below.
Can be used in some embodiments, so as to by the molecule for being combined from the instillation composition of these peptide linkers dissociationExample includes：Saline solution, the solution with slightly acidic pH, subalkaline pH, with different surfaces activating agent (i.e. poly- PyrusussuriensissAlcohol ester), the solution of EDTA or EGTA.In certain embodiments, the factor being dripped from the dissociation of the linker that starting is combinedBy depending on the bond strength of the factor-connection base complex, the bond strength is determined in turn beam body by dissociation constant.DissociationConstant can design the linker/peptide/fit to carry out by using the knowledge of the chemistry of amino acids of the interested factorModification.
One example of a kind of modified deposited part or wound insert is the Ca that can be originated with reference to wound or other modes2+And it is retained in into that of the wound site.In the early process of wound healing, Ca2+Ion is typically from cell officePortion is discharged into extracellular space.Believe resulting high Ca2+Concentration is involved many cell processes in wound healingThe positive effector of (such as adhesion, migration and differentiation).As a kind of high Ca2+Concentration is to need or beneficial for the wound bedWhen, instiled with a kind of solution containing a kind of chelating agen (for example, EDTA) or flushing can be used for destroying Ca2+Be bound to deposited part andIn being attached to the wound bed.
Another kind of factor for being used in conjunction with is transferrinss, i.e., a kind of blood plasma egg combined for ferrumIn vain.Have shown that chronic wounds fluid has significantly lower Transferrin, this indicates that oxidative stress occurs in chronic woundIn mouthful.It is known that free iron can work in the formation of free radical.Without being bound by theory, high-caliber free iron can be withCause the deterioration of tissue injury and the wound healing of delay.Transferrinss are combined and concentrated on and can be used for dissociating on deposited partFerrum is sequestered in wound bed, subsequently with a kind of appropriate instillation solution discharges together and subsequently after using negative pressure withExudate is removed together.This special time dependency regulation and control of transfer and iron level can provide a kind of notable benefit to patientPlace.One substitute viewpoint in, but transferrinss for the affinity of ferrum be it is very high be reversible because it withPH is reduced to progressively to reduce below neutrality.If being required to the needs of local concentration of iron, instil a kind of low ph solutionCan be used for from transferrinss releasing or discharging ferrum.
Hyaluronic acid (Hyaluronan) or hyaluronic acid (hyaluronic acid) are to be envisaged for together with the present inventionThe alternatively possible target for using.It is without being bound by theory, before being instiled for release with a kind of appropriate solution, will be saturatingBright matter acid is fixed and is concentrated to wound bed and can be used for promoting keratinocyte propagation and migrate, and reduces collagen deposition,Know this measure and then cause the cicatrization of reduction.Hyaluronic acid is also because of its free radical scavenging function it is well known that when it is tiedDuring the foam being bonded in wound location, the function can be beneficial.
During well-known lactoferrin is for embodiments of the invention because of antimicrobial and anti-inflammatory characteristics oneA little alternatively possible targets.There is synergism with FGF2 by the lactoferrin for having shown that endothelial cells secrete, becauseMigration of fibroblast cells and the ability of propagation is affected to dramatically increase together for them.It is specially designed it is fit can be used for combineBoth LF and FGF2, and discharge them in suitable treatment time scope with a kind of appropriate instillation solution.
TGFB-3 could be for the alternatively possible target of some in embodiments of the invention.This protein promoteesEnter the reorganization of substrate molecule, so as to cause the corium construction for improving and the cicatrization for reducing.TGFB-3 is to rise at itThe latent form for activating is needed to be secreted before effect.The activation of latency TGFB-3 via be bound to thrombin sensitivity eggIn vain -1 (TSP-1) occurs.Therefore, TSP-1 can in certain embodiments be used as a kind of composition in instillation fluid, to regulate and controlTGFB-3 is active.
Possible other targets also include calmodulin, CaM, S-100, thyroxine, Yi Jidan in addition to many otherHydrochlorate receptor.
C. the purposes of fit modified material
Modified polymer described above can be used for various purposes with the material being made from it, including (a) captureWith the biological target in concentration wound climate, (b) specify the chemical property for combining, and/or it is in be handed to that (c) specifies these target factorsThe orientation of cell.As discussed in more detail in examples section below, fit modified polyurethane (is modified including peptideGranuFoamTM(GF), i.e., a type of ROCF) can be used for capturing specified protein target in vivo.For example, peptide P16(a kind of anti-vegf peptide) and P22 (anti-GM-CSF peptide) are covalently bond to ROCF, and it is demonstrated that these fit capture target proteins.Example 2 and 3 provides the result of the amount of the GM-CSF for quantifying to be captured by the ROCF for being coated with P22, and example 4 is presented and quantified by applyingIt is covered with the amount of the VEGF of the ROCF captures of P16.
In certain embodiments, fit modified polymer can be used for combining the protein target in wound climate, andTheir signal transduction ability or other functions is not suppressed.This kind of fit modified polymer can serve as a kind of deposited part or woundA part for insert, for example, to capture and increasing the concentration of the biological activity protein through a wound bed of patient,For example, to strengthen or activate a kind of biological respinse of institute's targeting.
In certain embodiments, this deposited part can be used for interested protein such as VEGF collectionIn in wound bed, to cause angiogenesis and micro-pipe to be formed.Additionally, peptide can be designed to antagonism or chelate adversely shadowRing the protein of agglutination, such as matrix metalloproteinase.In certain embodiments, it is made up of this kind of fit modified polymerDeposited part therefore can be used for regulating and controlling different biological approach or for managing wound climate in unwanted biological activity pointThe presence of son or enzyme.
In addition to deposited part, fit modified polymer is (including using a kind of silyl-modified linker of isodigeranyl functionWith a kind of polymer based on polyurethane or copolymer those) can be also used for a lot of other materials, substrate and biological doctorIn learning device (including conduit).In this kind of embodiment, they can be used for conjugated various fit or other compounds, includingAntimicrobial.These materials are being discussed in further detail below to a kind of application of the therapy based on negative pressure.
The material of the present invention can also have advantages below：(either use with compound phase ratio well known in the prior artIn used in indication described herein or other), they can be more effectively, toxicity it is lower, more efficient, moreIt is virtuous, produce less side effect, more easily absorbed, and/or with more preferable phannacokinetic profile (exampleSuch as, higher oral bioavailability rate and/or lower clearance rate), and/or with other useful pharmacologys, physics or changeLearn characteristic.
It will be appreciated that the concrete anion or cation that form a part for any salt of the present invention are not crucial, onlyWill the salt be pharmacologically acceptable as an entirety.The other example and their system of pharmaceutically acceptable saltPreparation Method and purposes are presented on pharmaceutical salts handbook：In characteristics and uses (2002), the document is to be incorporated herein by reference.
D. the preparation of the insert comprising foamed materialss
On the other hand, the polymer based on foam can they are covalently attached to it is a kind of it is fit before or after enterRow is physically and/or chemically processed, coats or manipulated.Preparing some embodiments of modified wound insert includes：Compression (and/Or bonding) a kind of at least a portion of foam.Some embodiments include：Process and (for example, applied by applying heat or activationCoating) foam that compressed so that the foam is basic in the case where there is no external compressive force to keep being compressed.ExampleSuch as, in certain embodiments, processing includes being heated to being enough to the elastic rising for reducing the foam by the foam (for example, foam)Temperature.For example, the elasticity that the foam is heated to the foam can be lowered and/or is relaxed but less than the fusing of the foamOne temperature of temperature (for example, so that the foam will not be degraded by the elevated temperature).By this way, the foam can be withRelaxed the compression strain and compression produced in the foam using heat and pressure (compression stress).Generally, using high temperature realityThe now compression.In order to realize desired " deformation " so that the elasticity of the foam is lowered and/or the foam is not being depositedIt is basic in the case of compression stress to keep being compressed, temperature can be from the scope of 158 degrees Fahrenheits to 482 degrees Fahrenheits (for example, etc.In, less than, more than or anyone in the following between：140、160、180、200、220、240、260、280、300、320th, 340,360,380,400,420,440,460,480,500 degrees Fahrenheit, this depends on the concrete foam for being used).May be used alsoSo that foam is circulated to assist in keeping introduced deformation for example by a cooling, the foam can be cooled to less than room temperatureOr ambient temperature a temperature (for example, to or being equal to, less than, more than or anyone in the following between oneIndividual temperature：0th, 5,10,15,20,25,30,35,40,45,50,55,60,65 or 70 degrees Fahrenheit).It is a kind of in the formation of the present inventionIn some embodiments of the method for modified wound insert, by placement of foam in two plates for heating or platen (for example, in plateOr in pallet pressure machine and/or be wherein heated to these plates to be enough to the elastic temperature for reducing the foam) between；And activateThe forcing press so that these plates to be moved toward each other (for example, perpendicular to the thickness 320 of thickness portion 304) so that by the foamIntegral thickness or the degree of compression desired by being compressed to.This forcing press can electrically, machinery and/or hydraulic operation.
Some embodiments of the method for the wound insert that the manufacture of the present invention is modified also include：Cool down the foam (for example,After the foam is heated) so that, the compression section of the foam (for example exists in the case where there is no compression stress in room temperatureAt a temperature of 72 degrees Fahrenheits) under basic keep being compressed.In other embodiments, cool down the foam include cooling applied toThe coating of the foam so that the part compressed in the case where there is no compression stress being equal to, less than, more than or withIt is basic at the temperature between anyone or temperature range in lower items to keep being compressed：10、20、30、40、50、60、70、80、90th, 100,110,120,130,140, and/or 150 degrees Fahrenheit.
Can by any suitable method such as cut etc. to be formed in the foam in thick region and Bao QuDomain.This kind of method (such as) is taught by U.S. Patent Application Publication 2011/0178451, and the patent is incorporated by referenceHere.
In this kind of embodiment, the coating can be dispersed through foam, such as, by spraying the bubble with the coatingFoam, impregnates the foam, and/or any other the suitable mode for disperseing the coating in the foam in the coating.At someIn embodiment, for example, a transformation temperature at relatively low temperature (for example, less than single foam) can be occurred with apparatusDegree (for example, fusing point, glass transition etc.) produces a kind of material of rigidity coating the foam with being dried.In some enforcementsIn example, the coating may be configured to enable the foam compress (for example, without heating) at a lower temperature (and/or to pressCompression deformation) so that the coating becomes hard, or otherwise as coating is cooled down or is dried opposing expansion to keep the bubbleFoam is in its compressed configuration.For example, a kind of fluid adhesive can be applied to thickness portion before the foam is compressed andAllow to be dried before the compression stress is removed, so that the binding agent being dried will resist the expansion from the compressed thickness.At itIn his embodiment, the coating may be configured to make foam compression deformation so that the compression be it is reversible (for example, at leastIt is partially and/or fully reversible) so that the foam can expand and (for example, put with its heating or absorption moistureBe placed in a wound or on wound after).In certain embodiments, the coating comprising a kind of crosslinkable polymer and/orActivation includes exposing the coat to light and/or elevated temperature (for example, higher than ambient temperature, such as, it is sufficient to cause this canOne temperature of at least a portion crosslinking of the polymer of crosslinking), to cause at least certain part of the crosslinkable polymerBecome modified.
The example of suitable coating includes the crosslinkable polymer containing n- n-methylolacrylamide (NMA).NMA is canWith a kind of monomer with many other monomers (such as esters of acrylic acid and vinyl esters) copolymerization.In heating, (for example, to about 140DEG C), NMA contains group (such as carboxyl) reaction of hydroxyl with itself and other.Likewise it is possible to cause ureaformaldehyde, melamineThe polymer reaction that amine formaldehyde, and/or phenol formaldehyde contain hydroxyl with their own and other is forming crosslinking.Other cross-linking agent canTo include, for example, modified ethylene urea, the modified ethylene urea and the polymer containing hydroxyl react at elevated temperatures withThey are crosslinked.Other cross-linking agent can be included the peroxide of crosslinking most polymers at elevated temperatures.May be used alsoSo that the polymer containing hydroxyl and carboxylic group to be combined, and when heated, these polymer can form polyester friendshipConnection.In addition it is possible to use have at room temperature hypoergia and when heated can fast reaction with formed it is a kind of have hand overThe epoxy prepolymer of the epoxy polymer of connection.Likewise it is possible to using only at elevated temperatures and there is hydroxyl baseIn the case of group, amine or moisture just notable rapid-action polymeric isocyanate forming polyurethane or polyureas.
In certain embodiments, high-density region cooperates so as to being modified for the present invention with a kind of combination of density regionsWound insert provide different characteristic.For example, high-density region has a less aggregation hole size (aggregateCell size) and increased hole density, so high-density region has improved wicking function and more effectively transmitting streamBody (for example, more effectively detaches fluid and/or by from the fluid of a fluid source from the wound surface than density regionsUNICOM is to the wound surface).The mechanical strength of these high-density regions is also typically larger than these density regions, so that theseHigh-density region can provide structural support (example to these density regions and/or as overall modified wound insertSuch as, so that the modified wound insert is being not parallel to the side of these density regions upward against tear).In addition, thisA little density regions have a larger effective hole or pore size, so that these density regions are less susceptible to obstruction.EspeciallyIts, is when applying a negative pressure so as to from wound and when detaching fluid and/or exudate by the modified wound insert, theseThe larger pore size of density regions can allow fluid than aspirating fluid by the higher speed of these high-density regionsUnder be aspirated through these density regions so that microgranular and particulate material is sucked up to and/or by these low-densityRegion, to hinder and/or reduce these high-density regions in obstruction probability.In certain embodiments, it is also possible to use onePlant hydrophilic material to coat the foam to improve the wicking properties of be modified wound insert.
These density regions can be configured to allow the wound apply part bend and/or otherwise with oneWound is consistent.For example, these density regions can be relatively easily bent (and/or when by the modified wound insert edgeThe bending of density regions or there is relatively low elasticity when folding), to fold a modified wound insert, and/Or make a modified wound insert consistent with other hardware (such as plate, pin).
The typical foaming modifying wound insert of single density is isotropic, so that under negative pressure, a kind of typical caseThe foaming modifying wound insert of single density will proportionally shrink in all directions.In certain embodiments, the present inventionModified wound insert can be additionally configured to it is anisotropic so that the modified wound insert of the present invention can be withIt is configured to mechanically assist wound closure.For example, density regions it is lower than the density of high-density region (and under negative pressure willCompression is more).So, if applying negative pressure to modified wound insert, density regions will shrink than high-density regionIt is more, so that high-density region in the horizontal will shrink the wound insert for gathering together and being modified than in the verticalIt is more.In other embodiments, modified wound insert of the invention is configured with alternate and continuous largerClosed annular shape high-density region and density regions so that under negative pressure, the modified wound insert is by directionThe central cross of its own is to contract.
In certain embodiments, can coat and/or print thickness portion, thin section, high-density region and/or low density areaThe parent of foam of the domain (before the compression or after compression) to strengthen the individual region of the foam or as an entiretyWater or hydrophobic property.Other additives, such as antibiotic or drop blocking can also be contained and/or be coated with this kind of coated regionAgent (blockage-reducing agent).
In some embodiments of the invention, wound applies part comprising being configured to be positioned at a patient (for example, 30)One wound (for example, 26) on and/or the wound surface (for example, 42) on or a kind of wound for contacting with the wound surfaceDeposited part.
Some embodiments of the wound processing method of the present invention include：By a modified wound insert (for example, thisAny one in bright modified wound insert, such as 34a, 34b) it is positioned at a patient (for example a, wound 30)(for example, on 26), (for example, 404) and density regions (example the wherein modified wound insert comprising having high-density regionSuch as, 408) a kind of foam (density of the density that for example, 300), these density regions have less than these high-density regions.In certain embodiments, the foam is aseptic (for example, there is no microorganism and/or antibacterial).Some embodiments enter oneStep includes：One drop cloth (for example, 38) is attached to into the skin of neighbouring wound (for example, 46), so that the drop cloth covers this changesProperty wound insert and the wound, and between the drop cloth and the wound formed a space.Some embodiments include：It is logicalCross the deposited part (for example, by the modified wound insert) of the wound and apply negative pressure to the wound.In certain embodiments, willNegative pressure apply to the wound include actuation linkages to the wound apply part a vacuum source (for example, the equipment 14 of Fig. 1, or Fig. 3Vacuum source 200).Some embodiments include：Part is applied by the wound a kind of fluid is delivered to into the wound.In some embodimentsIn, delivering a kind of fluid includes that actuation linkages apply a fluid source (for example, the fluid source 248 of Fig. 3) of part to the wound.
Some embodiments of the wound treatment system of the present invention include (or the component of any embodiment of system 10 of system 10Any one subset) any one embodiment, and the present invention modified wound insert and/or wound apply part in oneIt is individual or multiple.
The different illustrative embodiments of device described herein, system and method are not intended to be restricted to disclosedThese concrete forms.But, they include all modifications and the replacement for falling within the scope of the appended claims.
Claim is not intended to include and be not necessarily to be construed as include that device plus function or step plus function formula are limited,Unless correspondingly using one or more terms " device being used for ... " or " step being used for ... in given claimSuddenly " clearly describing this restriction.
It should be appreciated that benefit described above and advantage can be related to one embodiment or can be related to several realitiesApply example.It will be further appreciated that, unless otherwise indicated, otherwise quote ' one ' project refer in those projects one or manyIt is individual.
The step of these methods described herein, can be carried out by any suitable order, or same in appropriate circumstancesShi Jinhang.
In the appropriate case, many aspects of any one example described above can be with any one describedThe many aspects combination of other examples, so as to formed with comparable to or different characteristic and address identical or notThe other example of same problem.
It should be appreciated that the explanation of above preferred embodiment is only given by way of example, and can be by this areaTechnical staff makes different modifications.Description above, example and data provide the structure and purposes of exemplary embodimentComplete description.Although below with a certain degree of concrete or with reference to one or more separate embodiments describe differenceEmbodiment, but those skilled in the art can do in the case of without departing substantially from the scope of the present invention to disclosed embodimentGo out a large amount of changes.
E. a kind of device of fit modified wound insert is included
Fit modified polymer described above can be used for various purposes with the material being made from it, including (a)Biological target in capture and concentration wound climate, is back to the wound bed, (b) chemical property that regulation is combined with optionally release,And/or (c) specifies these target factors in the orientation for being handed to these cells.
Can be with negative pressure wound therapy for a kind of deposited part or wound insert made by a kind of fit modified polymerIt is used together.In certain embodiments, being coated with fit or small peptide linker (part) compatible foam will whereby prevent themIn being removed to negative pressure wound therapy tank, when these are fit or beneficial molecule of the small peptide linker in wound fluid passes through the foamSelect them.Appropriate molecule for selecting from wound fluid includes flow of fluid will not be hindered to pass through the negative pressure wound therapyThe metabolite of deposited part, somatomedin, chemotactic factor and cytokine.In certain embodiments, appropriate linker will by withUnder it is this orientation combine these wound fluid molecules：" active site " of the wound fluid molecule is still available for for drawing one kindBiological respinse.For example, a kind of such molecule can be VEGF.VEGF has the special portion with reference to cell receptor on moleculePosition.The combination of the VEGF molecules and cell receptor can be used for a kind of initial biological respinse, typically angiogenesis.
In certain embodiments, can be used for combining in this these method taught and have chemotactic point to macrophageSon such as MCP1 (Figure 15), this can be used for stimulating expression of macrophage and migrates into wound and wound is advanced to from chronic more wherebyConjunction state.In certain embodiments, PDGF or collagen fragment can be combined in propagation or late phase inflammation phase process, to pierceSharp migration of fibroblast cells is into wound.Stimulating expression of macrophage and and then migration of fibroblast cells can help hinder into woundMouth progress is by inflammation phase and into the multiplicative stage of wound healing.In certain embodiments, nitric oxide synthetase canTo combine to stimulate perfusion from the wound fluid.This can aid in and enters the wound by allowing higher levels of nutrientAnd promote healing.In certain embodiments, can with reference to anti-inflammatory cytokine such as IL4 or IL10 to reduce inflammation, thereforeMake wound quickly be in progress by inflammation phase and enter the multiplicative stage of healing.DNA fragmentation can also be bound to into this to applyPart.In certain embodiments, the combination of highly charged DNA can make electric current pass through the deposited part.Electricity irritation is in woundMany years used in process.Therefore, DNA being bound to into the deposited part of the wound can allow to apply a current to the wound.
Tested for following：A kind of fit whether still may be used of peptide (a kind of antibody in this case) be bound toTo keep biological activity.The VEGF in solution is made by being connected to a kind of beadlet of VEGF antibody.Pass through this solution is madeAfter a little beadlet, centrifugation and these beadlet are washed.The beadlet of these washings is subsequently used for endothelial cell migration measure.ThisThe result of individual experiment shows after 3 hours, and compared with the beadlet without antibody, 2 times more of cell migration direction is bound to thisAntibody (and VEGF) these beadlet.Referring to Figure 18.It is therefore shown that being bound to a kind of VEGF of antibody can keep biological activity.
With reference now to accompanying drawing, and referring more specifically to Fig. 1, shown in it is the wound treatment system 10 of the present inventionOne of one embodiment.In an illustrated embodiment, equipment 10 includes a Wound treating apparatus 14 and by a conduitA kind of 22 wounds for being attached to equipment 14 apply part 18.(and connection is shown as indicated, applying part 18 and being configured to be attached toAn extremely) wound 26 of a patient 30.More specifically, in an illustrated embodiment, applying part 18 includes a kind of modified woundInsert 34 and a drop cloth 38.As indicated, modified wound insert 34 is configured to be positioned at (and is shown as positioning) on wound 26 (for example, in wound surface 42 or adjacent), and/or drop cloth 38 is configured to be attached to and (and is shownGo out to be attached to) skin 46 of the patient of neighbouring wound 26, so that drop cloth 38 covers the wound insert 34 that be modified and hindersMouth 26, and a space 50 is formed between drop cloth 38 and wound 26 (for example, wound surface 42).
Equipment 14 can include (for example) being configured to may be actuated (and/or actuating) so as to by negative pressure (for example, via conduit22) apply to wound apply part 18 a vacuum source, be configured to may be actuated (and/or actuating) so as to by a kind of fluid (for example,A kind of for example a kind of medicinal fluid of instillation fluid, antibacterium fluid, rinse fluid art etc.) delivering (for example, via conduit 22) to woundOne fluid source of deposited part 18.System 10 can be by any different configuration similar from those described in prior artAnd/or patient 30 is implemented and/or activated and/or be attached to method.For example, different wound therapy systems and component be by with/Or from Texas, USA, the KCI u s companys of San Antonio (San Antonio) and/or its subsidiary and correlationCompany's (being referred to as, " KCI ") is commercially available.
Conduit 22 can include single lumen catheter (for example, a vacuum source and/or a fluid source and equipment 14 itBetween switch), or multiple single lumen catheters or a multi-cavity catheter can be included so that (such as) can individually and/or whileFluid is delivered and/or can be applied negative pressure to wound and applies part 18 by ground.In addition, conduit 22 can include, for example, for applyingNegative pressure and/or first chamber of fluid delivering, and for being attached to one or more pressure transducers to sense in drop clothAt least one other chamber of the pressure or negative pressure between 38 and surface 42.In certain embodiments, conduit 22 can include manyIndividual chamber, for example, has a center cavity for applying negative pressure and/or fluid delivering such as in a single conduit, and withThe chambers of the heart it is adjacent or around center cavity arrange one or more peripheral cavities so that these peripheral cavities could be attached to a pressureForce transducer is to sense a pressure or negative pressure between drop cloth 38 and surface 42 (such as in space 50).These chambers canWith other chambers for being arranged to there is a center cavity and radially about arrange in the center cavity, or in other suitable peacesRow.These chambers can also be provided in single conduit.In an illustrated embodiment, system 10 further includes to be configured to connectionA wound for being connected to (and being shown coupled to) conduit 22 applies part connection gasket 54.One of a kind of suitable connection gasket 54Example is " V.A.C. commercially available from KCIPad ".An a kind of example of suitable drop cloth 38 includesCan it is commercially available from KCI "Drop cloth ".
With reference now to Fig. 2, it is shown that an a kind of side view of modified wound insert 34.Modified wound insert34 have a upside 100, downside 104, side 108,112 and internal volume 116.Although illustrate only modified woundThe side of mouth insert 34, but one of ordinary skill in the art will be understood that modified wound insert 34 includes one threeDimension rectangular volume, the volume has the depth vertically extended with shown side.In other embodiments, modified wound insertionThing 34 can have arbitrarily suitable shape, such as, cylinder form, imaginary shape, or can be rested and reorganized into unificationA kind of irregular shape of wound (for example, 26 and/or wound surface 42).Modified wound insert 34 can be comprising one kindFoam, such as, open-celled foam (it can also be netted).
The embodiment of the wound processing method of the present invention is may be better understood with reference to Fig. 1, system 10 is this diagram depictsOne schematic block diagrams of one embodiment.In an illustrated embodiment, wound applies part 18 and is attached to equipment 14, and setsStandby 14 include by a conduit 208 be attached to a tank 204 (for example, be configured to from wound apply part 18 receive exudate andOr the like) a vacuum source 200 (for example a, vacuum pump and/or analog).In an illustrated embodiment, equipment 14Further include：One pressure transducer 212, it has and is attached to conduit 208 by conduit 220 and/or three way cock 224One first pressure transducer 216；Change with the second pressure that tank 204 and/or the deposited part 18 of wound are attached to by conduit 232Can device 228.Pressure transducer 212 be configured to sense wound apply part 18 in and/or be attached to wound apply part 18, pressure sensingNegative pressure in any one of the different cavity (for example, in conduit) of device 212, and/or vacuum source 200.
In an illustrated embodiment, equipment 14 further includes a pressure relief valve 236 for being attached to conduit 232.ThisOutward, in an illustrated embodiment, tank 204 and vacuum source 200 are attached to wound and apply part 18 by conduit 240；And/or tank 204 canTo be included in the exit of tank 204 or a filter 244 of near exit, to prevent liquid or solid granule from entering conduit208.Filter 244 can be including being (for example) hydrophobicity and/or a lipophilic biofilter, so that aqueousAnd/or oil-based liquid will form liquid pearl on the surface of the filter.Equipment 14 is typically configured to so that in operating processIn, vacuum source 200 passes through a filter 244 by sufficient air-flow is provided so that the pressure drop on filter 244 is not notable(for example, so that the pressure drop substantive will not disturb the applying of to vacuum applying part 18 of the negative pressure from vacuum source 200).
In an illustrated embodiment, equipment 14 further includes to be attached to wound applies part 18 one by a conduit 252Individual fluid source 248, the conduit is such as attached to conduit 240 by a threeway or other suitable joints 256.In some realitiesIn applying example, three way cock 256 can include a switch valve and/or analog, so that can selectively allow in woundThe UNICOM applied between part 18 and fluid source 248 between deposited part 18 and vacuum source 200 or in wound.In certain embodiments, equipment14 only include one in vacuum source 200 and fluid source 248.In the embodiment that equipment 14 only includes fluid source 248, can also saveRemove tank 204 and/or pressure transducer 212.In different embodiments, as directed this embodiment, conduit 232 and/or conduit240 and/or conduit 252 can be combined and/or be included in a single multi-cavity catheter, as described above with reference to Figure 1.In certain embodiments, fluid source 248 be directly attached to wound apply part 18 (for example, one end of conduit 252 is such as via connection gasket54 are attached to wound applies part 18, and conduit 252 is attached on an opposite end fluid source 248；And conduit 252 is not attached to threePass joint 256).
In different embodiments, as shown in Figure 3 in this embodiment, equipment 14 be configured such that onceWhen liquid in the tank reaches a level of blocking filter 244, negative (or subatmospheric) pressure for greatly increasingPower will be produced in conduit 208 and sensed by transducer 216.Transducer 216 can be linked to circuit, and the circuit willThis pressure change is construed to the tank being filled and sends signal by means of the message and/or buzzer on a LCD to showTank 204 needs to be cleared and/or change, and/or automatically turns off or disable vacuum source 200.
Equipment 14 can be configured to by negative (or subatmospheric) pressure (for example, continuously, intermittently, and/orPeriodically) apply to the wound location, and/or cause pressure relief valve 236 to make the pressure energy in the wound site quickRecover to atmospheric pressure on ground.Therefore, if equipment 14 is programmed (such as) to discharge pressure with the interval of ten minutes, thenThese time intervals, pressure relief valve 236 can open the time limit of one section of regulation, it is allowed to which pressure is balanced in the wound site,And it is then shut off to recover negative pressure.It will be appreciated that when constant negative pressure is just being applied to the wound location, valve 236 is protectedClosure is held to prevent leakage into air or to prevent from atmospheric air leakage.In this state, it is possible in the wound site dimensionNegative pressure is held, without continuously operating and/or pump operation 200, and is only every now and then or periodically, to remain desiredPressure-reduction level (i.e. the desired pressure of subatmospheric power), the pressure-reduction level is sensed by transducer 216.Which save dynamicPower and enable facility use battery-powered operations longer period.
In certain embodiments, it is possible to use electric pulse, light, ultrasound and temperature are removing the factor or regulate and control theirsConcentration.
F. instil solution
In certain embodiments, by disclosed by here applying part made by these fit modified polymer can be with woundInstillation solution is used together, for example, applying a kind of Negative pressure into the wound of a patient.In certain embodiments, shouldInstillation solution includes the composition for being used to help discharge these factors for being bound to the foam or regulate and control its release.
Can be used for the example of the instillation composition in some embodiments so as to the combined molecule that dissociates includes：Salt is water-solubleLiquid, the solution with subacidity pH, the solution with alkalescence pH, with the molten of different surfaces activating agent (i.e. polysorbate)Liquid, the solution with slight ionic charge, EDTA or EGTA.In certain embodiments, the factor that starting is combined is from the connectionThe instillation fluid of the dissociation of base is by depending on the bond strength of the factor-connection base complex, the bond strength and then by the solutionDetermine from constant.The dissociation constant can count the connection by using the knowledge of the chemistry of amino acids of the interested factorBase/peptide is modifying.
In certain embodiments, the instillation solution includes hypochlorous acid (HOCl) and hypochlorite ion.Both it is used forThe example of the effective antimicrobial of biocidal effect.For example, HOCl be typically capable of kill wide spectrum microorganism (for example,Funguses, antibacterial, virus, funguses, yeast etc.)；Jing often (for example, can be in a section less than 10 seconds within the relatively short time periodThe microorganism more than 99% is killed in time).With fluid (for example, such antimicrobial can pass through the reactant of the present inventionWater and/or aqueous solution, such as, saline solution) combination produce or form, and can be than the institute in past treatment of woundsThe antimicrobial that the antibiotic for using is usually used with other is more effective and/or more general.For example, antibiotic can be thinBacterium-specificity, so that may require testing to determine a kind of suitable antibiotic for a kind of specific wound or infection；And/or so that antibiotic may only have the limited effectiveness for being directed to independent wound and/or infection (for example, not performingIn the case of test and/or in the case where a wound is by various different bacterium infections).This test may take up toThe time of several days, to determine a kind of appropriate antibiotic, treats or a kind of selection of effective antibiotic so as to delay.ThisOutward, antibacterial may develop the resistance to antibiotic, so that antibiotic may after a certain amount of time have reductionEffect.Additionally, antibiotic (systematically) gives typically in the way of vein, so that antibiotic may kill beneficialAntibacterial (for example, in the digestive system of patient) and/or organ injury (for example, to the liver of patient) may be caused.
By contrast, these reactants (and/or antimicrobial products of these reactants) of embodiments of the invention canThe wide spectrum for killing multiple-microorganism (for example, funguses, antibacterial, virus, funguses, yeast, etc.) is killed with being configured to haveAbility.Additionally, the reactant (and/or antimicrobial products of these reactants) of the present invention can be delivered partly (so as to anti-Only the systematicness of organ etc. is damaged or other side effect).
However, due to HOCl or OCl-With the reactivity of oxidable organic substance, its reality in wound care applicationWith being restricted before property.For example, producing hypochlorous art methods requires the electrolysis of saline etc. (for example, patient'sThe expensive equipment of bedside).Another example is lifted, commercially available chemicals (for example, bleach) have 5% or biggerHypochlorous acid concentration, this concentration it is too high and do not allow medical science use (for example, cytotoxicity being caused).Additionally, be adapted toUnder medical science concentration (for example, 2 to 20mM hypochlorite solutions), about 99% or more of the solution is water, so that shipment ratioIt is required more expensive and/or more difficult.Additionally, the storage of hypochlorite solution is difficult, because with the reaction of container typicallyDegrade or reduce the concentration of the hypochlorite solution.However, the wound insert of the present invention can deposit reactant (has depositionReactant in the foam of these wound inserts), so that when a kind of fluid such as saline or water is applied, OCl (and/Or ClO-) be released (for example, to form hypochlorous acid) and be delivered to a wound for biocidal effect.
G. example
Following instance is included to prove a preferred embodiment of the present invention.It will be understood by those of skill in the art thatThese technologies disclosed in these examples for below following are represented and are discovered by the present inventors Function in the practice of the inventionGood technology, and therefore be considered and constitute the preference pattern put into practice for it.However, in view of present disclosure, sheetThe technical staff in field in disclosed specific embodiment it should be appreciated that can make many changes, and still obtainSame or analogous result is without departing from the spirit and scope of the present invention.
The preparation of example 1-30 PHR shitosan boundary layers and silanization
In this program, a kind of isodigeranyl function silane crosslinker 3- amino-propyl diisopropyl Ethoxysilanes are sunkOn the shitosan foam of 30 parts of resin (30phr) parts per hundred parts, the foam is using the isocyanide of a kind of aromatic series two per 1.0g to productThe shitosan (Sigma (Sigma) production code member 448877) of acid esters and three branched chain polyol mixture 0.3g is producing.ThenBy multiple-OH groups of copolymer 3- aminopropyl diisopropyl Ethoxysilane (GelestTMCompound SIA0602.0)Carry out silylanizing.This is completed using scheme further below.
Determined resulting silicyl using a kind of o-phthalaldehyde(OPA) (OPA) for comparing Relative fluorescence units (RFU)The 30phr shitosans foam of change is compared with silylated reticulated, open type foam.OPA is bound to the primary-NH at it2BaseFluorescence is sent during group.Therefore, the RFU of 30phr shitosans foam should be more than (such as) OPA individually, so as to indicate the primary-NH2'sExist.The result of this test is illustrated in table 1 below and Fig. 5.This quantitative analyses confirm that the silylated 30phr shells gatherThe number (1.25e7) of the RFU that sugarwhips (" 30PHR Si ") have is the silylated reticulated, open type foam (ROCFSi) relative twice (6.45e6).This result has than the silylanizing with silylated 30phr shitosans foamThe more free amino group groups of reticulated, open type foam it is consistent.
The OPA measurement results of the PHR Si of table 1.30 and reticulated, open type foam (" ROCF ")
Fig. 5 is depicted to be determined using o-phthalaldehyde(OPA) (OPA) and is carried out the silylated 30phr shitosans foam of comparisonThe knot of (" 30PHR Si ") and the number of the free amino group group of silylated reticulated, open type foam (" ROCF Si ")Really.These results are provided according to Relative fluorescence units (RFU).OPA is individually the Sensitive Detection reagent for amine, and it also serves as rightAccording to.
Include the equipment used in these experiments is carried out：
Microplate reader (uncle rises (Biotek)-Synergy 4)
It is capable of the microscope (Olympus (Olympus)-IX51) of TRITC
One piece of 30phr shitosan foam
PBS bags (silent science and technology (the Thermo Scientific) 28384 of match)
96 holes, black, transparent bottom (Ge Laina (Greiner) 655209)
3- aminopropyl diisopropyl Ethoxysilanes, cover this specialization compound SIA0602.0 (" compound 602 ").ThisOne compound is kept drying.
25mL glass scintillation bottles
O-phthalaldehyde(OPA) or OPA (Sigma P/N-P0532)
1 inch of cork borer
4mm biopsy piercers
The test program for being used includes following steps：
Prepare 95% EtOH solution.
The 15%3- aminopropyl diisopropyl ethoxies in 95%EtOH are prepared in 25mL scintillation vials in fume hoodBase silane solution, and place it in and continue on agitator 5 minutes.
By wrapping PBS powder and 500mL DI H by 12O carries out mixing to prepare PBS.
Using a prepared food microtome from multiple thin slices of foam block cutting 2mm thick 30phr foams.
Sample is punched out with 1 inch of cork borer.
All samples preparation process is performed in glass scintillation bottle.
Sample is immersed in the 10mL bottles of 15% solution of silane and with a Glass rod relative to the bottleBottom is compressed, and to remove to be trapped in any bubble of the foam inside and be placed on an agitator 15 minutes are continued.
By being prepared in a similar manner negative control thing, except the 95%EtOH solution substituted silanes with 10mL it is moltenLiquid.
These samples are removed from the solution of silane with pliers and is transferred in 95%EtOH baths and is gently vortexedAbout 3 seconds to wash excessive silane off.Then this washing is repeated.
Then these samples are transferred in a clean 250mL beaker to guarantee that they will not contact with each other.
The beaker is placed on to be preheated in 80 DEG C of baking oven and continues 2 hours.After heating, by the beaker from thisBaking oven is removed and allows it to cool down at room temperature.Alternately, these samples can be placed in unlapped beaker simultaneouslyAnd allow to be incubated 24 hours at room temperature.
Using 4mm biopsies piercer from the multiple samples of silylated and not silylated 30phr punching presses.AlsoEach sample is weighed.
Quantization step：
ο adds the OPA of 200 μ L to each sample on the orifice plate of black 96, and the plate is covered and in room with paillon foilThe lower incubation of temperature 2 minutes.
The plate is placed in microplate reader ο and starting OPA determines scheme.
A kind of example 2-capture peptide is conjugated with the sulfo group-EMCS of 30PHR shitosan boundary layers
Parts per hundred parts the shitosan sample of 30 parts of resin (30phr) is the shitosan (Sigma P/N 448877) for using 0.3gPrepared with 1.0g polyurethane.Following test equipment, material and program are used to carry out these experiments.
Include the material used in these experiments is carried out：
The shitosan sample of 1 inch of silanization.
Sulfo group-EMCS (Pierre Si (Pierce) -22307)
BupH borate buffer bags (Pierre Si -28372)
EDTA (Sigma-Aldrich (Sigma Aldrich)-E9885-500G)
25mL scintillation vials.
Include the material used in these experiments is carried out：
The test program for being used includes these steps listed below：All buffer agents and reagent are all previously prepared, except sulfo group-EMCS, it is reconstructed immediately before the use.Referring to sulfo group-EMCS product description (Sai MokeSkill-Pierre Si), the description combines in full here by quoting with it.
1.1.BupH borate buffer solution：Description according to manufacturer adds in two bag BupH borate buffers powderTo 1L DI H2O。
1.2. conjugated buffer (BupH borate buffer solutions/5mM EDTA) is by the way that 146mg EDTA are dissolved inPrepare in 500mL BupH borate buffer solutions.
1.3. sulfo group-EMCS, it is allowed to be heated to room temperature, was carried out immediately before the sulfo group-EMCS addition steps are performedReconstruct.50mg sulfo group-EMCS are carried out weight by the description according to manufacturer in 1.22mL BupH borate buffer solutions/EDTAStructure is obtaining 100 × storing solution.
1.4. capturing peptide is reconstructed according to the description of manufacturer.Prepare 10mM working solutions.
1.5. sample preparation steps are performed in glass scintillation bottle so that the protein that adsorbed by this bottle and peptideAmount minimize.
1.6. general introduction.The experiment is carried out as follows (about 2.5 hours)：Reactant mixture 1 → washing → reactant mixture 2 →Washing → use or store at 4 DEG C.
1.8. reactant mixture 1:It is prepared by 100 × sulfo group-EMCS.This reaction is bound to the sulfo group-EMCS modifiedOn shitosan substrate.
1.8.1. the preparation of sulfo group-EMCS.BupH borate buffer solutions/the EDTA of 1.22mL is added to sulfo group-EMCS50mg bottles in and and then fully vibrate and be vortexed about 10 seconds.Then it is allowed to dissolve about 2 minutes, and 15It is used in the reconstruct of minute.
1.8.2. will be transferred in each bottle of reactant mixture from the shitosan sample of example 1.Negative control does not haveThere are sulfo group-EMCS or peptide；BupH borate buffer solutions/EDTA replaces these reagents in the reaction.
1.8.3. Glass rod is used to extrude the shitosan sample of these silanizations to guarantee the uniform of these foam surfacesReactivity.
1.8.4. 1 mixture of reaction is placed at room temperature and continues 30 minutes in an oscillator.
Once 1.9. the sulfo group-EMCS is conjugated, these samples are washed in BupH borate buffer solutions.Glass rod is used forThese bubbles are extruded into these samples to guarantee to remove all excessive sulfo group-EMCS.
1.10. the preparation of reactant mixture 2：Reactive polypeptide.
1.10.1. will add to the capture peptide (P22, commercially available anti-GM-CSF peptide) with reference to the foam of EMCS and incite somebody to actionBupH borate buffer solutions/EDTA mixture adds into each reaction bottle.
1.10.2. Glass rod is used to extrude these foam specimens to guarantee the uniform reactivity of these foam surfaces.
1.10.3. reactant mixture 2 placed on the oscillator at room temperature, continues 30 minutes.So by the capture peptideIt is conjugated to the sulfo group-EMCS cross-linking agent being deposited on the foam.
1.11. then these samples are washed in BupH borate buffer solutions to remove excessive peptide.Again, using dryNet Glass rod is removing these bubbles to guarantee uniform washing.
1.12. these samples are immediately available for into somatomedin and combine or these moist samples are transferred to into one to add a coverIn bottle, and them are stored at 4 DEG C until preparation is used, persistently less than 7 to 10 days.
1.13. by granulocyte macrophage colony stimulating factor (GM-CSF) GM-CSF of 333ng/mL and these samples onePlay incubation 24 hours.Wash these unconjugated protein off, then carried out with PBS buffer-exchanged and via ELISA quantifyBefore these samples are carried out into eluting with a kind of buffer (E1) based on imidazoles.These results indicate the positive sample with P-22Product capture the GM-CSF of 299pg/mL, and negative sample combines 124pg/mL.Referring to Fig. 8.
The capture of example 3-P22 modified open cell type network polymerss and CM-CSF
In this research, ROCF replaces the shitosan substrate described in the above example.DyLight Fluor moralsKe Sasi red (matching silent science and technology P/N 46412) is for replacing OPA.It is also a kind of NH2Chemically-reactive dyess；But, it is at oneFluorescence is sent under different wavelength.
This specialization compound 630 is covered for alternative cover this specialization compound SIA0602.0.Compound 630 has shorter alkaneChain and no propyl group, but carry out identical basic function.
It is bound to the fluorescence analysiss of-Sil of ROCF.In order to determine the silicyl or substituted first silicon that are bound on ROCFThe amount of alkyl group, by a kind of texas Red fluorescent dye (NH2Reactive fluorogen) it is conjugated be used for 630 linkers it is visualChange and quantify.630 isodigeranyl feature enables its Si atoms that the-OH groups on the foam are bound to an end,And another end is presented-a NH2Group, the group promotes NH2Reactive chemistry.Once this 630 is deposited on the ROCF, usePresence of the texas Red incubation identification cross-linking agent on the ROCF.Fluorescence analysiss show silicyl or substituted silicylGroup compound 630 is successfully deposited to ROCF (Fig. 9 A and 9B).The fluorescence of the test group quantifies to indicate to swash 584/630There are about 42,000 RFU under sending out/launching, and due to the background level of ROCF/ dyestuffs interaction, the negative control is (without firstThe ROCF of silylation or substituted silyl-group) produce 2,586.5 RFU of total.Therefore ,-Sil is conjugated causes about 16Fluorescence again increases, this number corresponding to the-Sil linkers being deposited on the foam.Also the ROCF in PBS and PBS is enteredGo and be analysed to ensure that the autofluorescence of these materials will not make RFU reading offsets.Data are given as meansigma methodss ± standard deviationGo out.
With-Sil the keys, then by " capture peptide " the conjugated ROCF to aminofunctional.For doing so, by example 2Sulfo group-EMCS the chemistry of development is used for commercially available GM-CSF antagonisies capture peptide P22 is conjugated to ROCF.ShouldP22- linkers-foam construct is incubated together with GM-CSF, strictly washing and eluting.By the GM-CSF of the elutingQuantified (Figure 11) via ELISA.The GM-CSF of the sample S3 many captures 49% more independent than ROCF with most P22.ByThe total amount (608pg/mL) of the GM-CSF of ROCF/P22 captures is more than being non-specifically bound to the negative control (ROCF is independent)GM-CSF amount (376pg/mL).Sample S1 to S3 is made up of P22- linkers-foam, wherein sulfo group-EMCS concentrationIncrease from S1 to S3.
The eluting result of P22/G-CSF experiments：P22 is obtained from the peptide catalogue of Bach's nurse u s company 2010.P22 be withA kind of inhibition mode combines the GM-CSF antagonisies of GM-CSF.Briefly, 330nmol P22 are chemical via sulfo group-EMCSIt is conjugated to ROCF.By P22- linkers-foam construct, overnight incubation, washing five times and are carried out together with 1 μ g GM-CSFStrict eluting.Washing before these eluting show the protein that non-specifically combines be reduced to insignificant level (<100pg/mL).In eluting, the sample S3 with most capture peptide is more than ROCF individually up to 49%.It is bound to ROCF/P22'sAmount (376pg/mL) of the total amount (608pg/mL) of GM-CSF more than the GM-CSF for being non-specifically bound to negative control C1.SampleProduct S1 to S3 is made up of peptide-linker-foam, and wherein the concentration of sulfo group-EMCS increases from S1 to S3.It is not available forThe positive control of this experiment, because also proving that peptide is covalently conjugated to polyurethane without technology.However, based on experiment knotReally, positive control is developed afterwards is used for subsequent experiment.Data are given as mean+/-standard error.
The capture of example 4-P16 modified polymer and VEGF
From the beginning P16 (a kind of anti-vegf peptide) is designed.The design is based on the dimeric dimer X-ray crystallography knots of VEGFStructure.Amino acid residue in actual biological design can be modified to strengthen or reducing binding affinity.
From the beginning P16 is designed.P16 is, based on the dimeric X-ray crystallography structures of VEGF, to deposit in Protein Data BankFor registration number l vpf.Showing naturally for dimerization of the generation between two VEGF homodimers is imitated in the P16 peptides designAs.
P16 is made up of a 11aa sequence of the dimer interface of the chain C in mimic protein data village, often used in village names structure l vpf.The P16 sequences are mainly made up of hydrophobic residue, the powered residue dispersed throughout of some of them.It is unique replace be at aa1, itsA kind of middle glycine replaces a kind of proline so that the steric hindrance farthest reduced from proline is disturbed.
Other experiment confirms P16 capture protein target VEGF.ELISA results from P16/VEGF experiments are indicatedP16 captures its protein target VEGF (Figure 14).By P16 it is conjugated be incubated to ROCF, with VEGF, strictly washing, soCarry out eluting with a kind of organic solvent afterwards.In this experiment, the test group (P16- linkers-foam) combines 584pg/mL,And maximum negative control combines 321pg/mL.Due to the hydrophobic property of both ROCF and target protein VEGF, it is contemplated that there is non-specificityWith reference to.However, these data indicate non-spy of the structure-based peptide design intensifier target specific protein capture beyond the ROCFDifferent in nature hydrophobic property.
These results summarized in both examples 3 and 4 show-Sil groups by successfully conjugated to ROCF.The positive is surveyedThe fluorescence of examination group quantifies instruction 21,702.5 RFU under 584/630 excitation/emission, and due to ROCF/ dyestuff phase interactionsBackground level, the negative control (ROCF without-Sil) is produced and amounts to 2,586.5 RFU.P16 and P22 is by successfully yokeIt is bonded to these linkers being deposited on the ROCF substrates.Finally, it is possible to quantify P16 and P22 and capture their target proteinEffectiveness.In the case of P16, elution profile indicates that positive is combined more than maximum negative control (ROCF of silanization)82% VEGF.Positive combines 584pg/mL, and the negative sample combines 321pg/mL.For P22, with maximumThe sample of capture peptide is more than ROCF individually up to 49%.The total amount (608pg/mL) for being bound to the GM-CSF of ROCF/P22 is more than non-spyThe amount (376pg/mL) of the GM-CSF of the negative control is bound to different in naturely.